ars.els-cdn.com€¦ · Web viewOne or two drops of conc. HCl was added in to sample tubes, 5 ml of ethyl acetate were added and vortex at high speed for 1 min and then the ethyl

Synergistic degradation of pyrene by five culturable bacteria in a mangrove sediment-derived

bacterial consortium

Pagakrong Wanapaisana, Natthariga Laothamteepa, Felipe Vejaranob, Joydeep Chakrabortyb,

Masaki Shintanic, Chanokporn Muangchindaa, Tomomi Moritab,d, Chiho Suzuki-Minakuchib,

Kengo Inouee, Hideaki Nojirib*, Onruthai Pinyakonga,f,g*

Department of Microbiology, Faculty of Science, Chulalongkorn University, 254 Phyathai

Road, Pathumwan, Bangkok 10330, Thailanda

Biotechnology Research Center, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo

113-8657, Japanb

Faculty of Engineering, Shizuoka University, 3-5-1 Johoku, Naka-ku, Hamamatsu, Shizuoka

432-8561, Japanc

Department of Bioscience and Engineering, Shibaura Institute of Technology, 307 Fukasaku,

Minuma-ku, Saitama, Saitama 337-8570, Japand

Department of Biochemistry and Applied Biosciences, Faculty of Agriculture, University of

Miyazaki, 1-1 Gakuenkibanadai-nishi, Miyazaki, Miyazaki 889-2192, Japane

Research Program on Remediation Technologies for petroleum contamination, Center of

Excellence on Hazardous Substance Management (HSM), Chulalongkorn University, 254

Phyathai Road, Pathumwan, Bangkok 10330, Thailandf

Omics Sciences and Bioinformatics Center, Faculty of Science, Chulalongkorn University,

254 Phyathai Road, Pathumwan, Bangkok 10330, Thailandg

*Address correspondence to Onruthai Pinyakong, [email protected] and to Hideaki

Nojiri, [email protected]

mailto:[email protected]

mailto:[email protected]

SUPPORTING INFORMATION Page

Extraction and analysis of pyrene and its intermediates. S1

Figure S1. Bacterial colony morphology on CFMM agar supplemented with 100

mg L-1 pyrene.

S2

Figure S2. Phylogenetic tree constructed based on 16S rRNA gene sequence. S3

Identification of putative RO genes S4

Figure S3. Dendrogram showing phylogenetic relatedness among reported PAH

dioxygenases.

S5

Table S1. Colony morphology of five bacterial strains on LB agar. S6

Table S2. Genome properties of draft genome sequences. S7

Table S3. Representative sequences belonging to each family of RO proteins. S8

Table S4. Closest biochemically studied RO identified for each homologue

obtained in this study and prediction of their putative substrate(s).

S10

Table S5. Putative genes involved in the degradation of pyrene and its

intermediate compounds.

S15

Table S6. Putative genes related to biosurfactant production. S30

Table S7. Degradation of protocatechuate (100 mg L-1) by strain FW1. S32

Table S8. The degradation rate of pyrene by bacteria. S33

Extraction and analysis of pyrene and its intermediates

One or two drops of conc. HCl was added in to sample tubes, 5 ml of ethyl acetate were

added and vortex at high speed for 1 min and then the ethyl acetate phase was removed into a

new tube. Then the sample was repeated the extraction with the ethyl acetate. After

dehydrating the extract by adding of anhydrous Na2SO4, the extract was evaporated to

remove all solvent. Methanol 1 ml was used to dissolve the compound. The dissolved sample

was filtered through a 0.2 μm PTFE filter prior to analyze by high performance liquid

chromatography (HPLC). HPLC was operated under the following conditions: an inertsill®

ODS column (4.6×150 mm) maintained at 40 °C; the mobile phase was 80% methanol; a

flow rate of 1.0 ml/min, and the UV detector was set at 275 nm.

The pyrene or intermediates degradation (%) was calculated as following:

(The quantity of pyrene or intermediates in control – The quantity of pyrene or intermediates in experiment) X 100The initial pyrene or intermediates in control

To estimate pyrene degradation rate (at day 3), the degradation rate was calculated as following:

Degradation rate = (Ct-C0)/(Tt-T0)

Where Ct is pyrene degradation at time t, C0 is pyrene degradation at time 0 and T is time.

The protocatechuate remaining (%) was calculated as following:

100 - (The quantity of protocatechuate in control – The quantity of protocatechuate in experiment) X 100The initial protocatechuate in control

S1

Figure S1. Bacterial colony morphology on CFMM agar supplemented with 100 mg L-1

pyrene

S2

PO1 PO2

Mycobacterium psychrotolerans MC6 (FJ770979) Mycobacterium psychrotolerans WA101T (AJ534886)

Mycobacterium psychrotolerans MCCC1A07723 (JX861600) Mycobacterium parafortuitum DSM 43528 (NR_026285)

Mycobacterium gilvum PYR-GCK (NC_009338) Mycobacterium vanbaalenii PYR-1 (NC_008726)

Mycobacterium sp. KMS (NC_008705) Mycobacterium sp. MCS (NC_008146) Mycobacterium sp. JLS (NC_009077)

FW1 Bacillus subtilis AER314-2 (KR967391) Bacillus subtilis TP-Snow-C17 (HQ327126) Bacillus licheniformis NIOT-ARMKVK06 (KC899772) Bacillus licheniformis AnBa3 (AY887128) Bacillus licheniformis MKU 8 (DQ071568)

Ochrobactrum intermedium NBRC 15820 (NR_113812) Ochrobactrum intermedium LMG 3301 (NR_026039)

PW1 Ochrobactrum anthropi BPyF3 (KJ782407) Ochrobactrum anthropi TCC-1 (KT820194) Novosphingobium naphthalenivorans TUT562 (NR_041046) Novosphingobium naphthalenivorans NBRC 102051 (NR_114027)

Novosphingobium pentaromativorans 17-33 (EU167957) Novosphingobium pentaromativorans 17-34 (EU167958) PY1 Novosphingobium pentaromativorans PQ-3 (KF145128) Novosphingobium pentaromativorans US6-1 (CP009291)

100

100

100

6299

100

100

100

8689

100

100

100

93

100

6888

76

7461

87

0.02

Figure S2. Phylogenetic tree constructed based on 16S rRNA gene sequence by neighbor-

joining methods. The numbers on the branch nodes indicate the percentages of bootstrap

support for the clades based on 1,000 bootstrap re-samplings.

S3

Identification of putative RO genes

In brief, 29 different ROs capable of transforming PAHs were identified from the RHObase

database. A careful inspection of the large (α) subunit sequence of RO terminal oxygenases

for the presence of conserved domains and a phylogenetic analysis (Figure S3) revealed that

all of the sequences belong to Class-A α3β3 type ROs [J. Chakraborty, D. Ghosal, A. Dutta, T.K.

Dutta, An insight into the origin and functional evolution of bacterial aromatic ring-

hydroxylating oxygenases. J. Biomol. Struct. Dyn. 30 (2012) 419–436.] and contain the

conserved RHO_alpha_C-NDO-like C-terminal domain [Conserved Domain Database, CDD

ID: cd08881]. Electron transport components (ETCs) are usually found to be associated with

these ROs were Rieske-type [2Fe-2S] ferredoxin, [3Fe-4S] ferredoxin, rubredoxin,

glutathione reductase (GR)-type and ferredoxin-NAD reductase fused with an N-terminal

plant-type [2Fe-2S] domain (FNRn). Thus, representative sequences belonging to each

above-described protein family were selected as queries for the BLAST search (Table S3).

S4

http://www.ncbi.nlm.nih.gov/pubmed/22694139

Figure S3: Dendrogram showing the phylogenetic relatedness among the reported PAH

dioxygenases. Abbreviations: αA, alpha subunit of Class-A ROs with a conserved

RHO_alpha_C_NDO-like C-terminal domain; β, beta subunit; Rsk, Rieske [2Fe2S]

ferredoxin; rub, rubredoxin; 3F4S, [3Fe-4S] ferredoxin; GR, glutathione reductase; and

FNRn, ferredoxin-NAD reductase fused with N-terminal plant-type [2Fe-2S] domain.

S5

Table S1. Colony morphology of five bacterial strains on LB agar

Bacterial strains Colony morphology

Mycobacterium sp. PO1

Shape: circularColor of colony: orange

Margin: entireElevation: raisedTexture: smooth

Mycobacterium sp. PO2

Shape: irregularColor of colony: orange

Margin: undulateElevation: flatTexture: rough

Novosphingobium pentaromativorans PY1

Shape: circularColor of colony: yellow


Ochrobactrum sp. PW1

Shape: circularColor of colony: cream


Bacillus sp. FW1

Shape: irregularColor of colony: white

Margin: undulateElevation: flatTexture: rough

S6

Table S2. Genome properties of draft genome sequences of strains isolated from pyrene-degrading consortium.

StrainGenome

size (Mb)Contigs Scaffoldsa

N50 with

scaffolds

Average

coveragebtRNA rRNA CDS GC(%)

Mycobacterium sp. PO1 5.83 183 153 76073 103 43 3 5574 68

Mycobacterium sp. PO2 5.83 186 158 77552 74 43 3 5583 68

Novosphingobium

pentaromativorans PY14.76 40 36 380690 183 49 3 4496 63

Ochrobactrum sp. PW1 4.74 47 32 453858 183 47 3 4566 58

Bacillus sp. FW1 4.06 20 18 507894 126 76 7 3970 44

a The number of scaffolds were estimated by the mapping of the paired reads against each of the assembled contigs.

b Based on the number of reads mapped to each contig, average coverage was calculated for the entire assembly of each strain.

S7

Table S3. Representative sequences belonging to each family of RO proteins which were

used as queries for blast search

Query a Accession no. Source organism

Nucleotide [gene] Protein

ALPHA SUBUNIT

Alpha-A

Alpha-B

Alpha-C

Alpha-Dcar

Alpha-Dksh

Alpha-Dvan

BphA1e

BphA1a

BphA1b

[PAH & alkylbenzene dioxygenase]

BphA1d [salicylate 1-hydroxylase]

BphA1c [salicylate 1-hydroxylase]

BphA1f [PAH dioxygenase]

XylX [Toluenebenzoate dioxygenase]

NidA3 [PAH dioxygenase]

PhtAa　[Phthalate dioxygenase]

AY365117 [nidA]

AF009224 [benA]

NC_008308 [andAc]

D89064 [carAa]

NC_000962 [kshA]

M22077 [vanA]

NC_002033

NC_002033

NC_002033

NC_002033

NC_002033

NC_002033

NC_002033

DQ028634

AY365117

AAT51751

AAC46436

YP_717961

BAA21728

NP_218043

AAA26019

NP_049179

NP_049184

NP_049186

NP_049206

NP_049213

NP_049062

NP_049209

AAY85176

AAQ91914

Mycobacterium vanbaalenii PYR-1

Acinetobacter sp. ADP1

Novosphingobium sp. KA1

Pseudomonas resinovorans CA10

Mycobacterium tuberculosis H37Rv

Pseudomonas sp. ATCC 19151

Novosphingobium aromaticivorans F199









FERREDOXIN

Rsk NC_002033 [bphA3] NP_049211 Novosphingobium aromaticivorans F199

Rub AF082663 [rub2] AAQ98844 Rhodococcus sp. NCIMB 12038

3F4S

Pl

BphA3

AY365117 [phtAc]

FJ708486 [dpaAc]

NC_002033

AAQ91918

ACO92630

NP_049211


Burkholderia sp. JS667


REDUCTASE

GR AY365117 [phtAd] AAQ91919 Mycobacterium vanbaalenii PYR-1

S8

Query a Accession no. Source organism

Nucleotide [gene] Protein

FNRn

FNRc

BphA4

M83949 [nahAa]

AF311437 [tsaB2]

NC_002033

AAA25900

AAK37997

NP_049182

Pseudomonas putida G7

Comamonas testosteroni T-2

Novosphingobium aromatics F199

a Abbreviations: Alpha-B, alpha subunit of Class-B ROs having conserved RHO_alpha_C_AntDO-like C-

terminal domain; Alpha-C, alpha subunit of Class-C ROs having conserved RHO_alpha_C_ahdA1c-like C-

terminal domain; Alpha-Dcar, alpha subunit of Class-D ROs, having conserved RHO_alpha_C_DMO-like C-

terminal domain and involved in degradation of carbazole and oxoquinoline; Alpha-Dksh, alpha subunit of

Class-D ROs involved in degradation of ketosteroids; Alpha-Dvan, alpha subunit of Class-D ROs, having

conserved RHO_alpha_C_DMO-like C-terminal domain and involved in degradation of vanillate, toluene

sulfonate and dicamba; Pl, Plant-type [2Fe2S] ferredoxin; FNRc, Ferredoxin-NAD reductase fused with C-

terminal plant type [2Fe-2S] domain.

S9

Table S4. Closest biochemically studied RO identified for each homologue obtained in this study and prediction of their putative substrate(s). Oxygenase designation

Position in the Contig

Most similar known Rieske oxygenase [GenBank accession]

% Protein identity Predicted substrate(s)

PO1_2_alpha 25343-24192 Aromatic oxygenase (GbcA) from Pseudomonas aeruginosa PAO1 [NP_254097]

31.4 Unknown, needs experimental data

PO1_9_alpha1 139155-137980

3-Ketosteroid 9alpha-hydroxylase (KshA) from Mycobacterium tuberculosis H37rv [NP_218043]

79.0 Ketosteroid like 4-androstadiene-3,17-dione

PO1_9_alpha2 158496-157321

3-Ketosteroid 9alpha-hydroxylase (KshA) from Rhodococcus jostii RHA1 [WP_011596916]


PO1_16_alpha 41080-39668 Aromatic oxygenase from Sphingomonas wittichii RW1 [WP_011951187]


PO1_21_alpha 81319-82356 3-Ketosteroid 9alpha-hydroxylase (KshA) from Rhodococcus opacus B-4 [BAH52700]


PO1_27_alpha 101431-100403

3-Ketosteroid 9alpha-hydroxylase (KshA) from Rhodococcus opacus B-4 [BAH52700]


PO1_31_alpha1Gene_1

(PO1_contig_31)*

119-1486 PAH dioxygenase (NidA) from Mycobacterium vanbaalenii PYR-1 [AAT51751]

99.6 Polycyclic aromatic hydrocarbons like naphthalene, phenanthrene, anthracene, benz[a]anthracene, fluoranthene, pyrene and benzo[a]pyrene;

Hetero polycyclic hydrocarbons like carbazole and dibenzothiophene;

Alkylbenzenes like toluene and xylenePO1_31_alpha2 4513-5694 Aromatic oxygenase (PdoX) from

Mycobacterium sp. S65 [AAQ12033]95.1 Carboxylated aromatics like salicylate, o-halobenzoate and

anthranilatePO1_31_alpha3 23730-24626 PAH dioxygenase (NidA3) from

Mycobacterium vanbaalenii PYR-1 [AAY85176] / Partial



Alkylbenzenes like toluene and xylenePO1_31_alpha4 30784-32199 Dibenzofuran dioxygenase (DfdA1) from

Terrabacter sp. YK3 [BAC06602]38.0 Hetero polycyclic hydrocarbons like dibenzofuran,

dibenzothiophene and 9-fluorenonePO1_31_alpha5 36297-37613 Dibenzofuran dioxygenase (DbfA1) from

Sphingomonas sp. KA1 [YP_718182]41.4 Hetero polycyclic hydrocarbons like dibenzofuran,

dibenzothiophene and 9-fluorenone

S10

Oxygenase designation




PO1_57_alphaGene_3

(PO1_contig_57)*

2231-3697 Phthalate 3,4-dioxygenase (PhtAa) from Mycobacterium vanbaalenii PYR-1 [AAQ91914]

95.3 Carboxylated aromatics like phthalate

PO1_58_alphaGene_2

(PO1_contig_58)*



PO1_74_alpha 104999-103656

Aromatic oxygenase from Mycobacterium avium subsp. paratuberculosis K-10 [WP_010949232]




PO1_102_alphaGene_2

(PO1_contig_102)*

989-2389 Phenanthrene 3,4-dioxygenase (PhdA) from Nocardioides sp. KP7 [BAA94708]



Alkylbenzenes like toluene and xylenePO1_116_alpha 4657-3371 2-Halobenzoate 1,2-dioxygenase (CbaA)

from Comamonas testosteroni BR60 [AAC45716]

35.6 Carboxylated aromatics like phthalate, chlorobenzoate, methoxy dichlorobenzoate, toluene-4-sulfonate, vanillate and phenoxybenzoate

PO2_1_alpha1 9472-10647 3-Ketosteroid 9alpha-hydroxylase (KshA) from Rhodococcus jostii RHA1 [WP_011596916]


PO2_1_alpha2 28813-29988 3-Ketosteroid 9alpha-hydroxylase (KshA) from Mycobacterium tuberculosis H37rv [NP_218043]




PO2_6_alpha1 2620-1304 Dibenzofuran dioxygenase (DbfA1) from Sphingomonas sp. KA1 [YP_718182]

41.4 Hetero polycyclic hydrocarbons like dibenzofuran, dibenzothiophene and 9-fluorenone

PO2_6_alpha2 8133-6718 Dibenzofuran dioxygenase (DfdA1) from Terrabacter sp. YK3 [BAC06602]

38.0 Hetero polycyclic hydrocarbons like Dibenzofuran, dibenzothiophene and 9-fluorenone

PO2_6_alpha3 15187-14291 PAH dioxygenase (NidA3) from Mycobacterium vanbaalenii PYR-1 [AAY85176] / Partial


S11






Alkylbenzenes like toluene and xylenePO2_6_alpha4 34404-33223 Aromatic oxygenase (PdoX) from

Mycobacterium sp. S65 [AAQ12033]95.1 Carboxylated aromatics like salicylate, o-halobenzoate and

anthranilatePO2_6_alpha5

Gene_39(PO2_contig_6)*

38798-37431 PAH dioxygenase (NidA) from Mycobacterium vanbaalenii PYR-1 [AAT51751]



Alkylbenzenes like toluene and xylenePO2_7_alpha 52991-54403 Aromatic oxygenase from Sphingomonas

wittichii RW1 [WP_011951187]37.7 Unknown, needs experimental data



PO2_53_alphaGene_14

(PO2_contig_53)*



PO2_55_alphaGene_2

(PO2_contig_55)*

989-2389 Phenanthrene 3,4-dioxygenase (PhdA) from Nocardioides sp. KP7 [BAA94708]

73.6 Polycyclic aromatic hydrocarbons like naphthalene, phenanthrene, anthracene, benz[a]anthracene, fluoranthene, pyrene and benzo[a]pyrene;Hetero polycyclic hydrocarbons like carbazole and dibenzothiophene;Alkylbenzenes like toluene and xylene

PO2_63_alpha 18345-19688 Aromatic oxygenase from Mycobacterium avium subsp. paratuberculosis K-10 [WP_010949232]


PO2_93_alpha 39538-40689 Aromatic oxygenase (GbcA) from Pseudomonas aeruginosa PAO1 [NP_254097]




PO2_108_alpha 2231-3697 Phthalate 3,4-dioxygenase (PhtAa) from Mycobacterium vanbaalenii PYR-1 [AAQ91914]

95.3 Carboxylated aromatics like Phthalate

PO2_116_alpha 73-1359 2-Halobenzoate 1,2-dioxygenase (CbaA) 35.6 Carboxylated aromatics like phthalate, chlorobenzoate, methoxy

S12





from Comamonas testosteroni BR60 [AAC45716]

dichlorobenzoate, toluene-4-sulfonate, vanillate and phenoxybenzoate

PY1_1_alpha1 389595-388381

Aromatic oxygenase from Hirschia baltica ATCC 49814 [WP_015828378]


PY1_1_alpha2 497722-496616

Dicamba O-demethylase (DdmC) from Stenotrophomonas maltophilia DI-6 [AAV53699]

36.0 Carboxylated aromatics like phthalate, chlorobenzoate, methoxy dichlorobenzoate, toluene-4-sulfonate, vanillate

PY1_4_alpha 286046-287242

Aromatic oxygenase from Parvibaculum lavamentivorans DS-1T [WP_012110685]


PY1_6_alpha1 148164-149249

3-Ketosteroid 9alpha-hydroxylase (KshA) from Rhodococcus opacus B-4 [BAH52700]

30.1 Ketosteroid like 4-androstadiene-3,17-dione;Hetero polycyclic hydrocarbons like carbazole and 2-

oxoquinolinePY1_6_alpha2 218346-

219548Aromatic oxygenase from Burkholderia

cenocepacia MC0-3 [WP_012337337]36.0 Unknown, needs experimental data

PY1_6_alpha3 219560-220900

Pre-Mupirocin oxygenase (MupW) from Pseudomonas fluorescens NCIMB 10586 [AAM12939]


PY1_6_alpha4 91413-92498 3-Ketosteroid 9alpha-hydroxylase (KshA) from Rhodococcus opacus B-4 [BAH52700]

50.4 Ketosteroid like 4-androstadiene-3,17-dione;Hetero polycyclic hydrocarbons like carbazole and 2-

oxoquinolinePY1_7_alpha1 215198-

214164Vanillate O-demethylase (VanA) from

Pseudomonas sp. HR199 [CAA72287]49.0 Carboxylated aromatics like phthalate, chlorobenzoate, methoxy

dichlorobenzoate, toluene-4-sulfonate, vanillate, phenoxybenzoate and mono- and di-chlorophenoxybenzoates

PY1_7_alpha2 264522-265895



PY1_9_alpha 271528-270158

Aromatic oxygenase from Sphingomonas wittichii RW1 [WP_011951187]


PY1_10_alpha1 13135-11702 Pre-Mupirocin oxygenase (MupW) from Pseudomonas fluorescens NCIMB 10586 [AAM12939]


PY1_10_alpha2 18918-17572 Aromatic oxygenase from Burkholderia cenocepacia MC0-3 [WP_012337337]




S13





PY1_10_alpha4 24244-25590 Aromatic oxygenase from Sphingomonas wittichii RW1 [WP_011951187]


PY1_10_alpha5 29115-27937 Aromatic oxygenase from Parvibaculum lavamentivorans DS-1T [WP_012110685]


PY1_10_alpha6 34548-35948 Pre-Mupirocin oxygenase (MupW) from Pseudomonas fluorescens NCIMB 10586 [AAM12939]


PY1_10_alpha7 35962-37146 Aromatic oxygenase from Parvibaculum lavamentivorans DS-1T [WP_012110685]


PY1_16_alpha?Gene_210

(PY1_contig_16)*

227268-228599

Phthalate 4,5-dioxygenase (Pht3) from Pseudomonas putida [BAA02511]

55.7 Carboxylated aromatics like phthalate, chlorobenzoate, methoxy dichlorobenzoate, toluene-4-sulfonate, vanillate, phenoxybenzoate and mono- and di-chlorophenoxybenzoates



PY1_17_alpha2 29029-30375 Aromatic oxygenase from Sphingomonas wittichii RW1 [WP_011951187]

27 Unknown, needs experimental data





PY1_17_alpha5 139602-138367

Aromatic oxygenase from Sphingomonas wittichii RW1 [WP_011951187]


PY1_17_alpha6 144986-146401



PW1_6_alpha 233224-231977

Aromatic oxygenase from Mesorhizobium loti MAFF303099 [WP_010912948]


Oxygenase designation is in the order – “strain name_contig number_oxygenase”*CDS number (contig number) referred in Table S4

S14

Table S5. Putative genes involved in the degradation of pyrene and its intermediate compounds.

CDS numbers(contig number)

Position

Top hit proteins by MiGAP Proteins in the reference strain

Predicted protein and its functionb

Name[GenBank accession]

Amino acid level identity (%)a

Namea[GenBank accession]


Mycobacterium sp. PO1 - Complete pyrene degradation

gene_1(PO1_contig_31) 119-1486

Ring hydroxilating dioxygenase subunit alpha of

Mycobacterium gilvum PYR-GCK [YP_001131939]

99.8

Ribosomal subunit interface protein of Mycobacterium

vanbaalenii PYR-1 [WP_011777803]

99.6

Pyrene hydroxilating dioxygenase α subunit (NidA). Similar to phenylpropionate dioxygenases and related ring-hydroxylating dioxygenases, large

terminal subunit

gene_20(PO1_contig_31) 19296-20756

NAD-dependent aldehyde dehydrogenase of

Mycobacterium sp. KMS [YP_937680]

96.7Aldehyde dehydrogenase of

M. vanbaalenii PYR-1 [WP_011777825]

96.5 2-Carboxybenzaldehyde dehydrogenase. NAD dependent aldehyde dehydrogenase

gene_44(PO1_contig_31) 40703-41482

Ribulose-5-phosphate 4-epimerase-like epimerase or aldolase of Mycobacterium

rhodesiae NBB3 [AEV73733]

99.2

3,4-Dihydroxyphthalate decarboxylase of M. vanbaalenii PYR-1

[ABM11387]

90.3 3,4-Dihydroxyphtalate 2-decarboxylase. Class II aldolase/adducing N-terminal

gene_43(PO1_contig_31) 41725-42576

2,3-dihydroxy-2,3-dihydrophenylpropionate

dehydrogenase of M. gilvum PYR-GCK [YP_001131898]

93.7

3-(cis-5,6-dihydroxycyclohexa-1,3-

dien-1-yl)propanoate dehydrogenase of M. vanbaalenii PYR-1 [WP_011777836 ]

96.2Dihydrodiol dehydrogenase. Dehydrogenases with

different specificities (related to short-chain alcohol dehydrogenases)

gene_3(PO1_contig_57) 2231-3697

Phthalate 3,4-dioxygenase alpha subunit (PadAa2) of Rhodococcus jostii RHA1

73.8Aromatic-ring-

hydroxylating dioxygenase subunit alpha of M.

96.5Phthalate 3,4-dioxygenase α subunit (PhtAa). Similar to phenylpropionate dioxygenase and related ring-hydroxylating dioxygenases, large

S15


Position







[YP_708559] vanbaalenii PYR-1 [WP_011777782] terminal subunit

gene_4(PO1_contig_57) 3730-4293

Phthalate 3,4-dioxygenase beta subunit (PadAb2) of R. jostii

RHA1[YP_708560]

68.3

3-Phenylpropionate dioxygenase of M. vanbaalenii PYR-1 [WP_011777783]

90.4 Phthalate 3,4-dioxygenase α subunit (PhtAb). Small subunit.

gene_6(PO1_contig_57) 4592-5395

2,3-Dihydroxy-2,3-dihydrophenylpropionate

dehydrogenase (PadB2) of R. jostii RHA1 [ YP_708562]

72



93.33,4-Dihydroxy-3,4-dihydrophtalate dehydrogenase (PhtB). Dehydrogenases with different specificities

(related to short-chain alcohol dehydrogenases)

gene_7(PO1_contig_57) 5424-5624

Phthalate 3,4-dioxygenase ferredoxin subunit (PadAc2) of R. jostii RHA1 [YP_708563]

60.6Ferredoxin of M.


89.4 Phtalate dioxygenase ferredoxin subunit (PhtAc)

gene_8(PO1_contig_57) 5621-6862

FAD-dependent pyridine nucleotide-disulfide

oxidoreductase of M. gilvum PYR-GCK [YP_001131868]

96.6

Pyridine nucleotide-disulfide oxidoreductase of


97.8 Phtalate dioxygenase reductase subunit (PhtAd). NAD(FAD)-dependent dehydrogenase

gene_11(PO1_contig_57) 8690-9775 Cupin of M. gilvum PYR-GCK

[YP_001131867] 95.6 Cupin of M. vanbaalenii PYR-1 [WP_011777787] 97.2 1-Hydroxy-2-naphthoate dioxygenase (PhdI).

gene_12(PO1_contig_57) 9794-10798

Dihydrodipicolinate synthetase of M. gilvum PYR-GCK

[YP_001131866]97.6 Aldolase of M. vanbaalenii

PYR-1 [WP_011777788] 97.6trans-2'-Carboxybenzalpyruvate hidratase-aldolase

(PhdJ). Dihydrodipicolinate synthase/N-acetylneuraminate lyase

gene_13 (PO1_contig_57) partially 10817-11119 First 100 a. a. of a

glyoxalase/bleomycin 99% in the homologou

First 100 a. a. of a glyoxalase of M.

98% in the homologous

Ring-cleavage dioxygenase (PhdF). Homologous to lactoylglutathione lyase and related lyases

S16


Position







assembledresistance protein/dioxygenase (296 a. a.) of M. gilvum PYR-

GCK [YP_001131865]s region vanbaalenii PYR-1

[WP_011777789] region

gene_2 (PO1_contig 58) 817-2283

Phthalate 3,4-dioxygenase alpha subunit (PadAa2) of Rhodococcus jostii RHA1

[YP_708559]

73.8

Aromatic-ring-hydroxylating dioxygenase

subunit alpha of M. vanbaalenii PYR-1 [WP_011777782]

95.1

Phthalate 3,4-dioxygenase α subunit (PhtAa). Similar to phenylpropionate dioxygenase and related ring-hydroxylating dioxygenases, large

terminal subunit

gene_3 (PO1_contig 58) 2280-2879


RHA1[YP_708560]

69.7


93.5 Phthalate 3,4-dioxygenase β subunit (PhtAb). Small subunit.

gene_5 (PO1_contig 58) 3178-3981



72.8



94.83,4-Dihydroxy-3,4-dihydrophtalate dehydrogenase (PhtB). Dehydrogenases with different specificities


gene_6 (PO1_contig 58) 4010-4210Phthalate 3,4-dioxygenase

ferredoxin subunit (PadAc2) of R. jostii RHA1 [YP_708563]




gene_7 (PO1_contig 58) 4207-5448



95.2




gene_8 (PO1_contig 58) 6026-7111 Cupin of M. gilvum PYR-GCK 99.7 Cupin of M. vanbaalenii 98.3 1-Hydroxy-2-naphthoate dioxygenase (PhdI).

S17


Position







[YP_001131867] PYR-1 [WP_011777787]

gene_9 (PO1_contig 58) 7130-8134Dihydrodipicolinate synthetase

of M. gilvum PYR-GCK [YP_001131866]

98.5 Aldolase of M. vanbaalenii PYR-1 [WP_011777788] 98.8

trans-2'-Carboxybenzalpyruvate hidratase-aldolase (PhdJ). Dihydrodipicolinate synthase/N-

acetylneuraminate lyase

gene_10 (PO1_contig 58) partially assembled 8153-8328

First 59 a. a. of a glyoxalase/bleomycin

resistance protein/dioxygenase (296 a. a.) of M. gilvum PYR-

GCK [YP_001131865]

100% in the homologou

s region

First 59 a. a. of a glyoxalase of M.


94.9% in the

homologous region

Ring-cleavage dioxygenase (PhdF). Homologous to lactoylglutathione lyase and related lyases

gene_99(PO1_contig_72)

101041-102186

Acetyl-CoA acyltransferase of Amycolatopsis mediterranei

U32 [YP_003766036]75.2

Acetyl-CoA acetyltransferase of M.


88.1 β-Ketoadipyl-CoA thiolase


Aromatic-ring-hydroxylating dioxygenase subunit beta M.

gilvum PYR-GCK [YP_001131852]

92.8


subunit beta of M. vanbaalenii PYR-1 [WP_011777802]

99.4Pyrene hydroxylating dioxygenase β subunit (NidB). Small subunit of ring-hydroxilating

dioxygenase

gene_3(PO1_contig_81) 2432-3229

Protocatechuate 3,4-dioxyenase subunit beta of Mycobacterium

sp. KMS [YP_937718]74.9

Protocatechuate 3,4-dioxygenase subunit beta of


80.2 Protocatechuate 3,4-dioxyenase β subunit

gene_4(PO1_contig_81) 3229-3792

Protocatechuate 3,4-dioxyenase subunit alpha of


72

Protocatechuate 3,4-dioxygenase subunit alpha of M. vanbaalenii PYR-1

[WP_011559101]

72 Protocatechuate 3,4-dioxyenase α subunit

S18


Position







gene_5(PO1_contig_81) 3789-4988

Fumarate lyase of Mycobacterium sp. KMS

[YP_937720]68.8

Fumarate lyase of M. vanbaalenii PYR-1 [WP_011777848]

69.1 β-Carboxy-cis,cis-muconate cycloisomerase. Adenylosuccinate lyase

gene_6(PO1_contig_81) 4985-6160

4-Carboxymucolactone decarboxylase of


70.73-Oxoadipate enol-lactonase

of M. vanbaalenii PYR-1 [WP_011777849]

71

γ-Carboxymuconolactone decarboxylase/ β-ketoadipate enol-lactone hydrolase. Homolog of gamma-carboxymuconolactone decarboxylase

subunit

gene_7(PO1_contig_81) 6157-68523-Oxoacid CoA-transferase subunit A of Mycobacterium

sp. KMS [YP_937722]82.8

3-oxoadipate CoA-transferase subunit A of M.


82.3β-Ketoadipate CoA-transferase α subunit. Acyl CoA:acetate/3-ketoacid CoA transferase, alpha

subunit

gene_8(PO1_contig_81) 6849-7532

3-Oxoacid CoA-transferase subunit B of Mycobacterium sp.

KMS [YP_951413]73.8

3-oxoadipate CoA-transferase subunit B of M.


73.8β-Ketoadipate CoA-transferase β subunit. Acyl CoA:acetate/3-ketoacid CoA transferase, beta

subunit


Aldehyde dehydrogenase of Mycobacterium sp. JLS

[YP_001069897]99.4

Aldehyde dehydrogenase of M. vanbaalenii PYR-1

[WP_011777801]99 Aldehyde dehydrogenase (NidD). NAD-dependent

aldehyde dehydrogenase

gene_4(PO1_contig_88) 4552-4043

Aromatic-ring-hydroxylating dioxygenase subunit beta of


100



99.4Pyrene hydroxylating dioxygenase β subunit (NidB2). Small subunit of ring-hydroxilating

dioxygenase

gene_2(PO1_contig_100) 144-1169


[YP_001131863]98.5

Dihydrodipicolinate synthetase of M. vanbaalenii

PYR-1 [WP_011777791]98.8 Hydratase-aldolase (PhdG). Dihydrodipicolinate

synthase/N-acetylneuraminate lyase

S19


Position









[YP_001131863]93.9




gene_2(PO1_contig_102) 989-2389

Ring hydroxylating dioxygenase subunit alpha Mycobacterium sp. KMS

[YP_937709]

99.1Ribosomal subunit interface

protein of M. vanbaalenii PYR-1 [WP_011777838]

99.6

Phenanthrene-4-carboxilate ring-hydroxylating dioxygenase α subunit. Similar to

phenylpropionate dioxygenases and related ring-hydroxylating dioxygenases, large terminal subunit

gene_3(PO1_contig_102) 2454-2972


Mycobacterium sp. JLS [YP_001069945]

100



100 Phenanthrene-4-carboxilate ring-hydroxylating dioxygenase β subunit. Small terminal subunit

Mycobacterium sp. PO2 - Complete pyrene degradation

gene_20(PO2_contig_6) 19621-18161

NAD-dependent aldehyde dehydrogenase of


96.7Aldehyde dehydrogenase of


96.5 2-Carboxybenzaldehyde dehydrogenase. NAD dependent aldehyde dehydrogenase

gene_39(PO2_contig_6) 38798-37431

Ring hydroxilating dioxygenase subunit alpha of

Mycobacterium gilvum PYR-GCK [YP_001131939]

99.8

Ribosomal subunit interface protein of Mycobacterium


99.6

Pyrene hydroxilating dioxygenase α subunit (NidA). Similar to phenylpropionate dioxygenases and related ring-hydroxylating dioxygenases, large

terminal subunit

gene_40(PO2_contig_6) 39404-38859

Aromatic-ring-hydroxylating dioxygenase subunit beta M.


99.4



92.8Pyrene hydroxylating dioxygenase β subunit (NidB). Small subunit of ring-hydroxilating

dioxygenase

S20


Position







gene_17(PO2_contig_24) 19804-20949Acetyl-CoA acyltransferase of Amycolatopsis mediterranei

U32 [YP_003766036]75.2





First 83 a. a. of an aromatic-ring-hydroxylating dioxygenase subunit beta (181 a. a.) of M.



s region

First 83 a. a. of an aromatic-ring-hydroxylating

dioxygenase subunit beta (169 a. a.) of M. vanbaalenii

PYR-1 [WP_011777800]

100% in the homologous

region

Pyrene hydroxylating dioxygenase β subunit (NidB2). Small subunit of ring-hydroxilating

dioxygenase

gene_4(PO2_contig_53) 3711-2686


[YP_001131863]98.5




gene_6(PO2_contig_53) 5201-4311

Glyoxalase/bleomycin resistance protein/dioxygenase

of M. gilvum PYR-GCK [YP_001131818]

98 Glyoxalase of M.


98.3 Ring-cleavage dioxygenase (PhdF). Homologous to lactoylglutathione lyase and related lyases

gene_7(PO2_contig_53) 6224-5220


[YP_001131866]98.5 Aldolase of M. vanbaalenii

PYR-1 [WP_011777788] 98.8trans-2'-Carboxybenzalpyruvate hidratase-aldolase

(PhdJ). Dihydrodipicolinate synthase/N-acetylneuraminate lyase

gene_8(PO2_contig_53) 7328-6243 Cupin of M. gilvum PYR-GCK

[YP_001131867] 99.7 Cupin of M. vanbaalenii PYR-1 [WP_011777787] 98.3 1-Hydroxy-2-naphthoate dioxygenase (PhdI).

gene_9(PO2_contig_53) 9147-7906



95.2




S21


Position







gene_10(PO2_contig_53) 9344-9144

Phthalate 3,4-dioxygenase ferredoxin subunit (PadAc2) of R. jostii RHA1 [YP_708563]




gene_11(PO2_contig_53) 10176-9373



71.6



943,4-Dihydroxy-3,4-dihydrophtalate dehydrogenase (PhtB). Dehydrogenases with different specificities


gene_13(PO2_contig_53) 11074-10475


RHA1[YP_708560]

69.7


93.5 Phthalate 3,4-dioxygenase α subunit (PhtAb). Small subunit.

gene_14(PO2_contig_53) 12537-11071

Phthalate 3,4-dioxygenase alpha subunit (PadAa2) of

Rhodococcus jostii RHA1[YP_708559]

73.8


subunit alpha of M. vanbaalenii PYR-1 [WP_011777782]

95.1

Phthalate 3,4-dioxygenase α subunit (phtAa). Similar to phenylpropionate dioxygenase and related ring-hydroxylating dioxygenases, large

terminal subunit


Ring hydroxylating dioxygenase subunit alpha Mycobacterium sp. KMS

[YP_937709]

99.1Ribosomal subunit interface

protein of M. vanbaalenii PYR-1 [WP_011777838]

99.6

Phenanthrene-4-carboxilate ring-hydroxylating dioxygenase α subunit. Similar to

phenylpropionate dioxygenases and related ring-hydroxylating dioxygenases, large terminal subunit

gene_3(PO2_contig_55) 2454-2972


Mycobacterium sp. JLS [YP_001069945]

100


subunit beta of M. vanbaalenii PYR-1

100 Phenanthrene-4-carboxilate ring-hydroxylating dioxygenase β subunit. Small terminal subunit

S22


Position







[WP_011559094]


Aldehyde dehydrogenase of Mycobacterium sp. JLS

[YP_001069897]99.4

Aldehyde dehydrogenase of M. vanbaalenii PYR-1

[WP_011777801]99 Aldehyde dehydrogenase (NidD). NAD-dependent

aldehyde dehydrogenase

gene_4(PO2_contig_77) 4321-4043

Last 92 a. a. of an aromatic-ring-hydroxylating dioxygenase

subunit beta (169 a. a.) of Mycobacterium sp. KMS

[YP_937656]


s region

Last 92 a. a. of an aromatic-ring-hydroxylating

dioxygenase subunit beta (169 a. a.) of M. vanbaalenii

PYR-1 [WP_011777800]

98.9% in the

homologous region

Pyrene hydroxylating dioxygenase β subunit (NidB2). Small subunit of ring-hydroxilating

dioxygenase

gene_7(PO2_contig_114) 4225-3542

3-Oxoacid CoA-transferase subunit B of Mycobacterium sp.

KMS [YP_951413]73.8




subunit

gene_8(PO2_contig_114) 4917-4222

3-Oxoacid CoA-transferase subunit A of Mycobacterium

sp. KMS [YP_937722]82.8




subunit

gene_9(PO2_contig_114) 6089-4914

4-Carboxymucolactone decarboxylase of


70.73-Oxoadipate enol-lactonase

of M. vanbaalenii PYR-1 [WP_011777849]

70.1

γ-Carboxymuconolactone decarboxylase/ β-ketoadipate enol-lactone hydrolase. Homolog of gamma-carboxymuconolactone decarboxylase

subunit

gene_10(PO2_contig_114) 7285-6086

Fumarate lyase of Mycobacterium sp. KMS

[YP_937720]68.8



S23


Position







gene_11(PO2_contig_114) 7845-7282

Protocatechuate 3,4-dioxyenase subunit alpha of


72


[WP_011559101]

72 Protocatechuate 3,4-dioxyenase α subunit

gene_12(PO2_contig_114) 8642-7845Protocatechuate 3,4-dioxyenase subunit beta of Mycobacterium

sp. KMS [YP_937718]74.9



80.2 Protocatechuate 3,4-dioxyenase β subunit

gene_5(PO2_contig_119) 1108-1887

Ribulose-5-phosphate 4-epimerase-like epimerase or aldolase of Mycobacterium

rhodesiae NBB3 [AEV73733]

99.2

3,4-Dihydroxyphthalate decarboxylase of M. vanbaalenii PYR-1

[ABM11387]

90.3 3,4-Dihydroxyphtalate 2-decarboxylase. Class II aldolase/adducin N-terminal

gene_4(PO2_contig_119) 2130-2981

2,3-dihydroxy-2,3-dihydrophenylpropionate

dehydrogenase of M. gilvum PYR-GCK [YP_001131898]

93.7



96.2Dihydrodiol dehydrogenase. Dehydrogenases with

different specificities (related to short-chain alcohol dehydrogenases)

Ochrobactrum sp. PW1 - Protocatechuate degradation

gene_205(PW1_contig_1)

221785-220721

3-Carboxy-cis,cis-muconate cycloisomerase of Brucella suis

1330 [YP_005614338]65.5




222406-221789

Protocatechuate 3,4-dioxygenase subunit alpha

(pcaG) of B. suis 1330 [YP_005614337]

85.4



25 Protocatechuate 3,4-dioxyenase β subunit

S24


Position








223148-222408

Protocatechuate 3,4-dioxygenase subunit beta

(pcaH) B. suis 1330 [YP_005614336]

93.1


[WP_011559101]

25.6 Protocatechuate 3,4-dioxyenase α subunit


223552-223145

4-Carboxymuconolactone decarboxylase (PcaC) of B. suis

1330 [YP_005614335]87.7

C-terminal domain of 3-Oxoadipate enol-lactonase of M. vanbaalenii PYR-1

[WP_011777849]

49.6 γ-Carboxymuconolactone decarboxylase


224358-223567

3-Oxoadipate enol-lactone hydrolase (PcaL) of B. suis

1330 [YP_005614334]73.4

N-terminal domain of 3-Oxoadipate enol-lactonase of M. vanbaalenii PYR-1

[WP_011777849]

32.1 β-ketoadipate enol-lactone hydrolase


228663-229370

3-Oxoadipate CoA-transferase subunit A (PcaI) of B. suis 1330

[YP_005614329]89.8




subunit


229367-230056

3-Oxoadipate CoA-transferase subunit B (PcaJ) of B. suis 1330

[YP_005614328]92.1




subunit

gene_216(PW1_contig_1) 230066-231268

Beta-ketoadipyl CoA thiolase of Ochrobactrum anthropi

ATCC 49188 [YP_001372253]91.2




Novosphingobium sp. PY1 - Phtalate degradation

S25


Position







gene_226(PY1_contig_7)

212816-211857

Oxidoreductase domain-containing protein of Delftia sp.

Cs1-4 [YP_004487731]70.7

Oxidoreductase of Novosphingobium

pentaromativorans US6-1 [WP_007014531]

78.7 2-Hydroxy-4-carboxymuconate semialdehyde hemiacetal dehydrogenase


213755-212847

Protocatechuate 4,5-dioxygenase of Delftia sp. Cs1-

4 [YP_004487058]58.3

Protocatechuate 4,5-dioxygenase subunit beta of N. pentaromativorans US6-

1 [WP_007014338]

28 Protocatechuate 4,5-dioxygenase β subunit


214167-213757

Protocatechuate 4,5-dioxygenase subunit alpha of

Delftia sp. Cs1-4 [YP_004488370]

57

Protocatechuate 4,5-dioxygenase subunit alpha of N. pentaromativorans US6-1 [WP_007014533]

97.8 Protocatechuate 4,5-dioxygenase α subunit


216228-215203

Amidohydrolase 2 of gamma proteobacterium NOR5-3

[ZP_05126814]70.1

4-oxalomesaconate hydratase of N.


97.1 4-oxalmesaconate hydratase


218457-219131

Methyltransferase (plasmid) of Sinorhizobium fredii NGR234

[YP_002823841]64

4-Carboxy-4-hydroxy-2-oxoadipate

aldolase/oxaloacetate decarboxylase of N.


98.7 4-hydroxy-4-methyl-2-oxoglutarate aldolase


219124-220188

Methylitaconate delta2-delta3-isomerase of Novosphingobium

sp. PP1Y [YP_004538832]99.4

4-oxalomesaconate tautomerase of N.


99.7 4-Oxalomesaconate tautomerase

S26


Position








220181-221071

2-pyrone-4,6-dicarbaxylate hydrolase of Novosphingobium

sp. PP1Y99.3

2-Pyrone-4,6-dicarboxylate hydrolase of N.


99 2-Pyrone-4,6-dicarbaxylate hydrolase


223185-222346

Protocatechuate 4,5-dioxygenase subunit beta of Novosphingobium sp. PP1Y

[YP_004538829]

98.6


1 [WP_007014338]

29.3 Protocatechuate 4,5-dioxygenase β subunit

gene_238(PY1_contig_7) 223600-223190

Protocatechuate 3,4-dioxygenase of

Novosphingobium sp. PP1Y [WP_041558615]

83.8




225920-224928

4,5-Dihydroxyphthalate decarboxylase of Burkholderia sp. RPE64 [YP_008040264]

64.2

4,5-Dihydroxyphthalate decarboxylase of

Burkholderia cepacia DBO1 [AAD03553]

40.3 4,5-Dihydroxyphthalate decarboxilase (OphC)


227119-225917

1-Carboxy-3-chloro-3,4-dihydroxycyclo hexa-1,5-diene

dehydrogenase of Brucella melitensis bv. 1 str. 16M

[NP_541843]

24.8Phthalate dihydrodiol dehydrogenase of B.

cepacia DBO1 [AAD03557]45.9 cis-Phthalate dihydrodiol dehydrogenase (OphB)


227268-228599

Vanillate O-demethylase oxygenase of Caulobacter

crescentus CB15 [NP_421196]20.5 Phthalate dioxygenase of B.

cepacia DBO1 [AAD03558] 52.2 Phthalate 4,5-dioxygenase (OphA2)

S27


Position








125522-124611

Protocatechuate 4,5-dioxygenase of Delftia sp. Cs1-

4 [YP_004487058]56.2


1 [WP_007014338]

25.3 Protocatechuate 4,5-dioxygenase β subunit


125932-125522

Protocatechuate 4,5-dioxygenase subunit alpha of

Delftia sp. Cs1-4 [YP_004487729]

51.6



a. Percent identities calculated from pair-wise alignments using EMBOSS Stretcher. For partially assembled CDS, EMBOSS Water as also used.b. Following: Kim S, Kweon O, Jones RC, Freeman JP, Edmondson RD, Cerniglia CE. 2007. Complete and integrated pyrene degradation pathway in

Mycobacterium vanbaalenii PYR-1 based on systems biology. J Bacteriol 189(2):464-472. Kim S, Kweon O, Jones RC, Edmondson RD, Cerniglia CE. 2008. Genomic analysis of polycyclic aromatic hydrocarbon degradation in Mycobacterium vanbaalenii PYR-1. Biodegradation 19:859-881. Chang HK, Zylstra GJ. 1998. Novel organization of the genes for phthalate degradation from Burkholderia cepacia DBO1. J Bacteriol 180(24):6529-6537.

Luo YR, Kang SG, Kim SJ, Kim MR, Li N, Lee JH, Kwon KK. 2012. Genome sequence of benzo(a)pyrene-degrading bacterium Novosphingobium pentaromativorans US6-1. J Bacteriol 194(4):907.

S28

Table S6. Putative genes related to biosurfactant production.

CDS numbers(contig number) Position

Top hit proteins by MiGAP Proteins in the reference strains

Predicted function of the proteinb





gene_234(FW1_contig_7)

227642-228316

4'-phosphopantetheinyl transferase of Bacillus sp. JS

[YP_006230168]100

Biosurfactant biosynthesis protein

(Sfp) of Bacillus licheniformis NIOT-

AMKV06[AGR39386]

95.5

Activation of peptidyl carrier protein (PCP) domains of surfactin

synthetases

gene_239 (FW1_contig_7)

232660-233388

Surfactin synthetase of Bacillus sp. JS [YP_006230163] 100

Biosurfactant biosynthesis protein (SrfA) of Bacillus

licheniformis NIOT-AMKV06

[AGR39388]

96.7

Thiosterase domain of surfactin synthetases,

responsible for cyclisation of surfactin

peptide


233417-237244

Surfactin synthetase of Bacillus sp. JS [YP_006230162] 100

SfrA3 of Bacillus subtilis JH642

derivative W168 [CAA49818]

94.7

Non-ribosomal peptide synthetase modules. In charge of recognition, activation and covalent

binding of residues in the

S29

CDS numbers(contig number) Position

Top hit proteins by MiGAP Proteins in the reference strains

Predicted function of the proteinb





surfactin peptide chain


237281-248032

Surfactin synthetase of Bacillus sp. JS [YP_006230161] 99.2



95.1


248045-258808

Surfactin synthetase of Bacillus sp. QB928 [YP_006628533] 96.3



94

a. Percent identities calculated from global pair-wise alignments using EMBOSS stretcher.b. Following: Lawarence A, Balakrishnan M, Joseph TC, Sukumaran DP, Valsalan VN, Gopal D, Ramalingam K. 2014 Functional and molecular

characterization of a lipopeptide surfactant from marine sponge-associated eubacteria Bacillus lichenifromis NIOT-AMKV06 of Andaman and Nicobar Islands, India. Mar Poll Bull 82:76-85. Cosmina P, Rodriguez F, Ferra F, Grandi G, Perego M, Gerard V, Sinderen D. 1993. Sequence and analysis of the genetic locus responsible for surfactin synthesis in Bacillus subitlis. Mol Microbiol 8(5): 821-831.

S30

S31

Table S7. Degradation of protocatechuate (100 mg L-1) by strain FW1 in 5 ml of CFMM incubated at 200 rpm and at 30oC.

Strain Protocatechuate degradation (%)Day 2 Day 3 Day 4 Day 5

FW1 13.21 ± 1.60 18.64 ± 4.45 25.80 ± 4.38 26.48 ± 0.07

S32

Table S8. The degradation rate of pyrene by bacteria.

Bacterial strains or consortia Degradation rate (% day-1)

PO1 9.92PO2 9.19PY1 0.61PW1 1.92FW1 1.98P1: PO1+PY1 13.87P2: PO1+PW1 18.08P3: PO1+FW1 18.70P4: PO1+PO2+PY1 12.23P5: PO1+PO2+PW1 15.12P6: PO1+PO2+FW1 18.01P7: PO1+PO2 16.14C1: PO1+PO2+PY1+PW1+FW1 29.20C2: PO2+PY1+PW1+FW1 25.86C3: PO1+PY1+PW1+FW1 25.65C4: PO1+PO2 +PW1+FW1 28.92C5: PO1+PO2+PY1+ FW1 31.40C6: PO1+PO2+PY1+PW1 29.24C7: PY1+PW1+FW1 1.91

S33

Documents

ars.els-cdn.com€¦ · Web viewOne or two drops of conc. HCl was added in to sample tubes, 5 ml of ethyl acetate were added and vortex at high speed for 1 min and then the ethyl