Bacterial Polysaccharides - Welcome to the Complex ...rcarlson/Bacps.pdf · Polysaccharide...

Bacterial PolysaccharidesBacterial PolysaccharidesAnimal PathogensAnimal Pathogens•• Salmonella Salmonella enteriditisenteriditis••E. coliE. coli••NeisseriaNeisseria meningitidismeningitidis••HaemophilusHaemophilus influenzaeinfluenzae••Proteus mirabilisProteus mirabilis••Proteus Proteus vulgarisvulgaris••MycobacteriaMycobacteria(many others)(many others)

Soil BacteriaSoil Bacteria•• RhizobiaceaeRhizobiaceae••Pseudomonas Pseudomonas solanacearumsolanacearum••ErwiniaErwinia amylovoraamylovora(many others)(many others)

PolysaccharidesPolysaccharides••ExtracellularExtracellular polysaccharides (polysaccharides (EPSsEPSs))

••Excreted Excreted heteropolysaccharidesheteropolysaccharides••Most often acidicMost often acidic••Frequently form solutions of high viscosityFrequently form solutions of high viscosity••GramGram--positive and Grampositive and Gram--negative productsnegative products

••Capsular Polysaccharides (Capsular Polysaccharides (CPSsCPSs))••CellCell--associated associated heteropolysaccharidesheteropolysaccharides••Most often acidic, but some are neutralMost often acidic, but some are neutral••Solutions can be highly viscousSolutions can be highly viscous••GramGram--positive and Grampositive and Gram--negative productsnegative products

••LipopolysaccharidesLipopolysaccharides ((LPSsLPSs))••Component of the outer membraneComponent of the outer membrane••AmphiphilicAmphiphilic••GramGram--negative bacterial componentnegative bacterial component

••TeichoicTeichoic acidacid••GramGram--PositivePositive

••Mycobacterium cell wall componentsMycobacterium cell wall components••MycolylMycolyl arabinogalactanarabinogalactan••LipooligosaccharidesLipooligosaccharides ((LOSsLOSs))••GlycopeptidolipidsGlycopeptidolipids ((GPLsGPLs))••PhenolicPhenolic glycolipidsglycolipids ((PGLsPGLs))

Polysaccharide FunctionsPolysaccharide Functions

••Prevention of Prevention of desicationdesication••SurvivialSurvivial

••AdherenceAdherence••Colonization of oral surfacesColonization of oral surfaces••Colonization of cathetersColonization of catheters••BacteriaBacteria--plant interactionsplant interactions

••Resistance to nonResistance to non--specific host immunityspecific host immunity••ComplementComplement--mediated mediated phagocytosisphagocytosis••ComplementComplement--mediated killingmediated killing

••Resistance to specific host immunityResistance to specific host immunity••Poor antibody responsePoor antibody response

••CellCell--cell recognition cell recognition signallingsignalling••BacteriaBacteria--plant interactionsplant interactions

GramGram--Negative Bacterial Cell WallNegative Bacterial Cell Wall

CPS

EPS

E. coliE. coli Capsular PolysaccharideCapsular Polysaccharide

E. coli K30 wt

E. coli K30CPS minus

E. coli K30CPS/LPS minus

From"Current Topics in Microbiology and Immunology", 1990, Ed. by K. Jann and B. Jann

Examples of Capsular Polysaccharides fromExamples of Capsular Polysaccharides fromEnteric BacteriaEnteric Bacteria

44--ββ--DD--GlcAGlcA--1 41 4--αα--DD--GlcNAcGlcNAc--1 61 6--αα--DD--GlcNAcGlcNAc--11 nnLL--SerineSerine--6 6 (amide)(amide)

E. coliE. coli K40 CPSK40 CPS

22--αα--DD--ManMan--1 31 3--ββ--DD--GalGal--11 nnββ--DD--GlcAGlcA--1 31 3--αα--DD--GlcGlc--33

E. coliE. coli K30 CPSK30 CPS

Grou

p I

Grou

p I

Grou

p II

Grou

p II 88--αα--Neu5AcNeu5Ac--2 92 9--αα--Neu5AcNeu5Ac--22 nn

E. coliE. coli K92 CPSK92 CPS

33--ββ--DD--RibRib--1 71 7--ββ--KdoKdo--22 nnE. coliE. coli K23 CPSK23 CPS

(Whitfield and Valvano. 1993. Biosynthesis and expression(Whitfield and Valvano. 1993. Biosynthesis and expressionof cellof cell--surface polysaccharides in gramsurface polysaccharides in gram--negative bacteria. negative bacteria. Adv. Microbial. Adv. Microbial. PhysiolPhysiol..35:13535:135--246).246).

LipopolysaccharidesLipopolysaccharides: Structure, Synthesis, : Structure, Synthesis, FunctionFunction

READ: Raetz, and Whitfield. 2002. READ: Raetz, and Whitfield. 2002. LipopolysaccharideLipopolysaccharide EndotoxinsEndotoxins. Ann. Rev. . Ann. Rev. BiochemBiochem. 71:635. 71:635--700.700.

Schematic Diagram of a Schematic Diagram of a LipopolysaccharideLipopolysaccharide

Lipid ACore

Lipid ACoreO-Chain

Lipid ACoreO-Chain

LPS II

LPS I

LPS III, IV, V

Lipid A BiosynthesisLipid A Biosynthesis

Raetz, and Whitfield. 2002. Raetz, and Whitfield. 2002. LipopolysaccharideLipopolysaccharide EndotoxinsEndotoxins. Ann. Rev. . Ann. Rev. BiochemBiochem. 71:635. 71:635--700.700.

Lipid A and the Innate Immune ResponseLipid A and the Innate Immune Response

Raetz, and Whitfield. 2002. Raetz, and Whitfield. 2002. LipopolysaccharideLipopolysaccharide EndotoxinsEndotoxins. Ann. Rev. . Ann. Rev. BiochemBiochem. 71:635. 71:635--700.700.

Unusual Lipid A StructuresUnusual Lipid A Structures

Raetz, and Whitfield. 2002. Raetz, and Whitfield. 2002. LipopolysaccharideLipopolysaccharide EndotoxinsEndotoxins. Ann. Rev. . Ann. Rev. BiochemBiochem. 71:635. 71:635--700.700.

RhizobialRhizobial and and E. coliE. coli Lipid ALipid A

Rhizobiaceae

R. leguminosarumR. etli

O

OHHO

HO CO2 HCore

OHO

O

O

NHO

O O

O NHCO2 H

OHHO

CH2 O

OHO

HOOO

O

O

HO O

Rhizobium Lipid AE. coli Lipid A

(U.R. Bhat, L.S. Forsberg, & R.W. Carlson. 1994.J. Biol. Chem. 269:14402-14410; N. L. Que, S. Lin, R.J.Cotter & C.R. Raetz.2000. J. Biol. Chem. 275:28006-28016; N.L. Que, A.A. Ribeiro & C.R. Raetz2000. J. Biol. Chem. 275:28017-28027)

OO

O

NHO

O OHO

ONH

O

O

P

P

O

O

HO

OH

OH

OH

OHO

O

HO

O

O

O O

HO

((BrucellaBrucella, , BartonellaBartonella))

Biosynthesis of Unusual Lipid A StructuresBiosynthesis of Unusual Lipid A Structures

Raetz, and Whitfield. 2002. Raetz, and Whitfield. 2002. LipopolysaccharideLipopolysaccharide EndotoxinsEndotoxins. Ann. Rev. . Ann. Rev. BiochemBiochem. 71:635. 71:635--700.700.

O

NHO

-O3POO

O

HO

O

NHO

HOOPO3

-

O

HO CH2OH

CO2H

HO

O

O

HO

HO CH2OH

CO2H

HO

O

O

HOO

HO

O

HO

UDP-GlcNAc1

1. 3-O-acylation (lpxA).2. De-N-acetylation (lpxC).3. N-acylation (lpxD).4. Disaccharide synthase (lpxB).5. 4’-phosphorylation (kinase).6. Kdo transferase (kdtA).

1. Price, Kelly, Raetz, and Carlson. 1994. J. Bacteriol.176:4646-4655.

2. Price, Jeyaretnam, Carlson, Kadrmas, and Raetz. 1995.PNAS 92:7352-7356.

3. Brozek, Kadrmas, and Raetz. 1996. JBC 271:32112-32118.

1-phosphatase34’-phosphatase2

27O

HC

28:0

Acp

-XL

4

4. Brozek, Carlson, and Raetz. 1996. JBC 271:32126-32136.

Man

LpcC5

5. Kadrmas, Allaway, Studholme, Sullivan, Ronson, Poole, and Raetz. 1998. JBC 273:26432-26440.

GalLpcA5

KdoLpcB5

GalA ?

GalA ?

GalA ?

GalA?

Glc

Nox

idat

ion7

(?)

acyl

ase6

6. Basu, White, Que, and Raetz. 1999. JBC 274:11150-11158.

7. N. L. S. Que, S. H. Lin, R. J. Cotter, and C. R. H. Raetz. 2000.JBC 275 (36):28006-28016.

Biosynthesis ofRhizobial LPS

Transport of Lipid ATransport of Lipid A

Raetz, and Whitfield. 2002. Raetz, and Whitfield. 2002. LipopolysaccharideLipopolysaccharide EndotoxinsEndotoxins. Ann. Rev. . Ann. Rev. BiochemBiochem. 71:635. 71:635--700.700.

Transport of Lipid ATransport of Lipid A

Raetz, and Whitfield. 2002. Raetz, and Whitfield. 2002. LipopolysaccharideLipopolysaccharide EndotoxinsEndotoxins. Ann. Rev. . Ann. Rev. BiochemBiochem. 71:635. 71:635--700.700.

Lipid A Core StructuresLipid A Core Structures

Schematic Diagram of a Schematic Diagram of a LipopolysaccharideLipopolysaccharide

Lipid ACore

Lipid ACoreO-Chain

Lipid ACoreO-Chain

LPS II

LPS I

LPS III, IV, V

Lipid A Core StructuresLipid A Core Structures

Raetz, and Whitfield. 2002. Raetz, and Whitfield. 2002. LipopolysaccharideLipopolysaccharide EndotoxinsEndotoxins. Ann. Rev. . Ann. Rev. BiochemBiochem. 71:635. 71:635--700.700.

Core BiosynthesisCore Biosynthesis

Raetz, and Whitfield. 2002. Raetz, and Whitfield. 2002. LipopolysaccharideLipopolysaccharide EndotoxinsEndotoxins. Ann. Rev. . Ann. Rev. BiochemBiochem. 71:635. 71:635--700.700.

Polymerization and Polymerization and LigationLigation of Oof O--Chain PolysaccharideChain Polysaccharide

n

0123456

Pare

nt

rfc

mut

ant

rfaL

mut

ant

Num

ber o

f rep

eat u

nits

(Figure taken from Whitfield. 1995. Biosynthesis of (Figure taken from Whitfield. 1995. Biosynthesis of lipopolysaccharidelipopolysaccharide O antigens.O antigens.Trends in MicrobiologyTrends in Microbiology 3:1783:178--185)185)

LipopolysaccharidesLipopolysaccharides: Structure, Synthesis, : Structure, Synthesis, FunctionFunction

Raetz, and Whitfield. 2002. Raetz, and Whitfield. 2002. LipopolysaccharideLipopolysaccharide EndotoxinsEndotoxins. Ann. Rev. . Ann. Rev. BiochemBiochem. 71:635. 71:635--700.700.

Rhizobium leguminosarum/etli LPS Core

O-Chain

O

O

CO 2H

O

CH 2O HO H

Lipid A

CO 2H

H O CH 2O H

OO

CO 2H

O

O

O

O

CO 2H

O

O

CO 2H

OH O

CH 2O HHO

O CO 2H

O

O

O

O H

H O

O H

O H

O H

O H

O H

O H

O H

H O

H O

H O

H O

O H

Kdo

Gal

Man

KdoGalA

GalA

GalA

Kdo

(U.R. Bhat, S.K. Bhagyalakshmi, & R.W. Carlson. 1991. Carbohydr. Res. 220:219-227,L.S. Forsgberg, & R.W. Carlson. 1998. J. Biol. Chem. 273:2747-2757)

LipopolysaccharidesLipopolysaccharides: Structure, Synthesis, : Structure, Synthesis, FunctionFunction

Raetz, and Whitfield. 2002. Raetz, and Whitfield. 2002. LipopolysaccharideLipopolysaccharide EndotoxinsEndotoxins. Ann. Rev. . Ann. Rev. BiochemBiochem. 71:635. 71:635--700.700.

LipopolysaccharidesLipopolysaccharides: Structure, Synthesis, : Structure, Synthesis, FunctionFunction

Raetz, and Whitfield. 2002. Raetz, and Whitfield. 2002. LipopolysaccharideLipopolysaccharide EndotoxinsEndotoxins. Ann. Rev. . Ann. Rev. BiochemBiochem. 71:635. 71:635--700.700.

LipopolysaccharidesLipopolysaccharides: Structure, Synthesis, : Structure, Synthesis, FunctionFunction

Raetz, and Whitfield. 2002. Raetz, and Whitfield. 2002. LipopolysaccharideLipopolysaccharide EndotoxinsEndotoxins. Ann. Rev. . Ann. Rev. BiochemBiochem. 71:635. 71:635--700.700.

Regulation of Polysaccharide Chain LengthRegulation of Polysaccharide Chain Length

rfb

rol

rfb

rol

rfb

rol

+

+

+

1 kb1 kb3 kb

orf1 orf2 orf3 wzm wzt

FAJ1

201

FAJ1

202

FAJ1

203

FAJ1

200

395

367

Ace

tyl

Ace

tyl t

rans

fera

setr

ansf

eras

e

Ace

tyl

Ace

tyl t

rans

fera

setr

ansf

eras

e

met

hyl

met

hyl t

rans

fera

setr

ansf

eras

e

met

hyl

met

hyl t

rans

fera

setr

ansf

eras

e

lpe3

lpeA

Fuco

seFu

cose

synt

hesis

synt

hesis

ABC

tra

nspo

rter

ABC

tra

nspo

rter

5

OO CH 3

O CH 3O H

H 3 C

O

CH 3

O

5 O-Acetyl groups

O HH OH O

O

OOO

H O

O HCO

O CH 3

O

O

H 3 C

O CH 3

H 3 C O

O

O H

O HO H

OO H

H 3 C

OH O

N H Ac

H O CH 3

OO

O

H OCH 2 O H

CO O H

H O

O

Core Lipid A

CH3

CH3

CH3

Kdo

QuiNAcManFuc

Fuc

dTal

GlcA

Fuc

25% 10%

367367

395395FAJ1201

FAJ1202

FAJ1203

FAJ1200

LipopolysaccharidesLipopolysaccharides: Structure, Synthesis, : Structure, Synthesis, FunctionFunction

Raetz, and Whitfield. 2002. Raetz, and Whitfield. 2002. LipopolysaccharideLipopolysaccharide EndotoxinsEndotoxins. Ann. Rev. . Ann. Rev. BiochemBiochem. 71:635. 71:635--700.700.