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BIOL710: Molecular BiologyMondays and Wednesdays, 5:30PM
LECTURERS
Mitchell GoldfarbHN834 772-5289Goldfarb@genectr.hunter.cuny.edu
Thomas Schmidt-GlenewinkelHN841 772-5027Thomas@genectr.hunter.cuny.edu
COURSE WEBSITE (for Goldfarb lectures)
http://biology.hunter.cuny.edu/Goldfarb/
Then click link to course at bottom of the web page
SUGGESTED TEXT
Biochemistry, 6th editionBerg, Tymockzo, & Stryer
ISBN: 0716787245OR
Biochemistry, 4th editionStryer
ISBN: 0716720094
Available online fromwww.barnesandnoble.com
www.amazon.com
Scope of the Course
How are biological macromolecules synthesized and assembled?How do different macromolecules generate the structure of cells?How do proteins fold to acquire functionality?How do enzymes catalyze reactions?How is energy harvested and stored in the cell?How do pumps and channels store energy and control the chemical composition of cellular compartments?Intracellular biochemical signaling by proteins and lipids.DNA structure and replication.Transcription and post-transcriptional RNA modification.Regulation of transcription.The genetic code and protein translation.Similarities and differences in processing of genetic information in prokaryotes vs. eukaryotes
Exams and Studying
Midterm on Oct. 22nd (tentative)Covers Lectures 1-12 on Proteins (Goldfarb and Schmidt-Glnewinkel)
Final in DecemberCovers Lectures 13-26 on DNA/RNA & Information Flow (Schmidt-Glenewinkel)
Study Tips
Read suggested readings before and/or soon after each lecture
Distribute your studying throughout semester; do not cram
LECTURE 1: pH, pK, Amino Acids, Peptide Properties, pIReading: Berg Chapters 1,2
Before being able to discuss the properties of biological macromolecules, we must first review:
1) Covalent and noncovalent bonding
2) Oxidation and reduction
3) Acid/base chemistry
Elemental Properties
Hydrogen
Carbon
Oxygen
Nitrogen
Sulphur
Phosphorus
H
C
O
N
S
P
ElectropositiveForms 1 covalent bond
Forms 4 covalent bonds
Strongly electronegativeForms 2 covalent bonds
Weakly electronegativeForms 3 covalent bonds
Weakly electronegativeForms 2 covalent bonds
In oxidized formForms 5 covalent bonds
CH
H
C
H
H
H
O H
CH
H
C
H
O
O
H
ethanol
acetic acid
CN
C
C
H
O
O
H
H
H
glycine
CN
H
C
H
O
O
H
H
H
HH
S Hcysteine
R -- O -- P -- O -- H
O
O- phosphate side group
Oxidation and Reduction of Organic Compounds and Oxygen
OXIDATION -- Removal of an electron pair from a moleculeREDUCTION -- Addition of an electron pair from a molecule
In any oxidation/reduction reaction, one component (electron donor) gets oxidized, another component (electron acceptor) gets reduced
CH
H
C
H
H
H
O H
CH
H
C
H
H
O2 H+2 e +
-
ETHANOL (reduced) ACETALDEHYDE (oxidized)
O2 + 2NADH + 2H+ 2H2O + 2NAD+
O
O
OH H
ACCEPTOR DONOR REDUCED OXIDIZED
Weak Acids and Bases
When hydrogen is covalently bonded to an electronegative atom(typically O or N) in a compound, the hydrogen proton may dissociate.
In this situation:The protonated compound is called an ACID,
and the deprotonated compound is called a BASE.
CH
H
C
H
O
O
H CH
H
C
H
O
O
-
H+
CO
H
C
H
H
N
H
H H
H
H
+
H+
CO
H
C
H
H
N
H
H
H
H
Conjugate ACID Conjugate BASE
acetic acid acetate
2-amino-ethanolhydroxyethylammonium
pH
Water is in equilibrium with its minor dissociation products, H+ and OH-
While concentration of H2O is 55 M, [H+][OH-] = 10-14 M
In a neutral solution, [H+] = [OH-] = 10-7 M
In an acidic solution, [H+] >> [OH-] . E.g. [H+] = 10-3 M , and [OH-] = 10-11 M)
In a basic solution, [H+] << [OH-] . E.g. [H+] = 10-10 M , and [OH-] = 10-4 M)
pH = - log10 [H+]
At neutrality, pH = - log10 [10-7] = 7Acidic solutions have pH < 7; Basic solutions have pH > 7
Generally speaking “weakly acidic” means pH = 4 to 5.5
“strongly acidic” means pH < 3“weakly basic” means pH = 8.5 to 10
“strongly basic” means pH > 11
Strong Acids and Bases
A strong ACID is an INORGANIC compound that FULLY DISSOCIATES into
H+ and its negative counterion in water.
A strong BASE is an METAL HYDROXIDE compound that FULLY DISSOCIATES into
OH- and its positive counterion in water.
pH of strong acid and base solutions
HYDROCHLORIC ACID NITRIC ACID
HCl H+ Cl- HNO3 NO3-H+
SODIUM HYDROXIDE CALCIUM HYDROXIDE
NaOH Na+OH- Ca(OH)2 Ca+2 2OH-
pH = - log10 [ACID] pH = 14 + log10 [BASE]
pH of 10 mM HCl = 2 pH of 0.1 M NaOH = 13
e.g. e.g.
TITRATION OF A STRONG ACID SOLUTION WITH STRONG BASE
14
0
0
2
2
1
1
54
3
876
1312
11109
NEUTRAL
pH
Molar Equivalents of NaOH
Different amounts of NaOH added to a 10mM HCl solution
pH stays strongly acidic until nearly 10mM NaOH added,then swings steeply past neutral to strongly basic when NaOH exceeds 10mM
NO CONTROL OF pH IN THE MILD ACID TO MILD BASE RANGE
pKa of an acid/base conjugate pair
HA H+ + A-Acid Proton Base
Ka = [H+ ][A- ]
[HA]
TAKE LOGARHYTHMMULTIPLY BY -1 pH = pKa + log10
[A-][HA]
IMPLICATIONS FOR ANY PARTICULAR ACID/BASE PAIR
For a group with an acidic pKa, the base conjugate predominates at neutral pH
For a group with an basic pKa, the acid conjugate predominates at neutral pH
When acid and base are at equal concentrations, pH = pKa
An acid/base pair acts as a buffer of strong acids or bases
in pH range of pKa + 1.
Titration of a Weak Acid Solution with a Strong BaseThe weak acid BUFFERsBUFFERs the effect of the strong base,
keeping pH in the range of the pKa over wide range of base concentratiion
As NaOH is added to a weak acid solution HA, the NaOH converts HA to A-
HA + NaOH H2O + Na+ + A-
Since pH = pKa + log [ A- ]/[ HA ] , the weak acid buffers in the range of pKa
0
0
2
1
1
0.5
5
4
3
7
6
Molar Equivalents of NaOH
pH
pH
For acetic acidpKa = 4.0
For unbuffered0.1 M acetic acid
pH = 2.5WHY???
pKa
25
75
100
50
[HA
] (mM
)[H
A] (m
M)
[A[A--] (m
M)
] (mM
)
pH of a Weak Acid Solution in Water
HA H+ + A-
pH = pKa + log10
[A-][HA]
For a weak acid in water: [A-] = [H+] and [HA] = [A]total
= pKa + log10 [H+] - log10 [A]total
Therefore:
pH = (pKa + log10 [A]total)
2
Three Types of Non-Covalent Bonding InfluenceIntramolecular and Intermolecular Interactions
ELECTROSTATIC BONDING
HYDROGEN BONDING
Oppositely charged groups are attractive.
R
C O
O- R
H
H
HN +
A hydrogen covalently bonded to O or N can noncovalently interact with a O or N, if all three atoms are aligned and at appropriate distance. This is a hydrogen bond.
R O H O
C
R
N H
H
O
H
R
OH
H
OH H
O
H
H
Three Types of Non-Covalent Bonding InfluenceIntramolecular and Intermolecular Interactions
VAN DER WAAL’S FORCES AND HYDROPHOBIC INTERACTIONS
A weak interaction between nonpolar molecular surfaces.Van der Waal’s forces contribute to favorability of hydrophobic interactions.The other crucial contributing factor is that interaction between two hydrophobic surfaces in a solution reduces the hydrophobic surface area and therby INCREASES the number of water-to-water solvent hydrogen bonds!!!
AMINO ACIDS
pK of carboxylic acid group is ~ 3.0pK of amino group is ~ 9.5
POLYPEPTIDES
FREEROTATING
FREEROTATING
All polypeptides synthesizedfrom L-amino acids
CLASSES OF AMINO ACIDS
All proteins are synthesized from a pool of 20 amino acids(some additional amino acids are generated by modifications within synthesized polypeptides)
Amino acids can be functionally grouped by properties of side chains (R)(a few amino acids fit overlap into more than one group)
GROUPINGS
ALIPHATIC -- Side chains participate in hydrophobic interactions
AROMATIC -- Hydrophobic interactions and hydrogen bonding
ACIDIC -- Ionic and hydrogen bonding
BASIC -- Ionic and hydrogen bonding
HYDROXYL -- Hydrogen bonding and sites of phosphate or sugar modifications
AMIDO -- Hydrogen bonding and sites of sugar modifications
SULPHUR -- Hydrogen bonding and sites of oxidative crosslinking
“OTHER” -- Side chains confer specialized turning properties of polypeptide
ALIPHATIC AMINO ACIDS
AROMATIC AMINO ACIDS
NOTE:
Tyrosine isalso a
hydroxylatedaminoacid
NOTE:
Methionine isalso a
sulphur-bearing
amino acid
HYDROXYLATED AMINO ACIDS
Ser-, Thr-, or Tyr-OH in protein can undergo phosphate addition
Ser- or Thr-OH in protein can can be site for carbohydrate addition(termed O-linked glycosylation)
ACIDIC AMINO ACIDS
AMIDO AMINO ACIDS
Carboxylic acid side chain in Asp and Gluhas pKa ~ 4.5 and carries
a full -1 charge at neutral pH
Asn is amidated version of AspGln is amidated version of Gln
Asn and Gln are NOT charged,but are higly polar
NH2 group on Gln in proteins can be sitefor carbohydrate addition(N-linked glycosylation)
BASIC AMINO ACIDS
Amino side chain of Lys and imino side chain of Arg have pK > 10and carry a full +1 charge at neutral pH
Ring imino side chain of His has pK ~ 6.5And carries on average a fractional positive charge at neutral pH
SULPHUR-CONTAINING AMINO ACIDS
Methionine is also consideredan aliphatic amino acid
2 H+2 e +
-Nearby cysteines on the
same or differentpolypeptide chains
can undergo oxidationto generate a covalent
DISULPHIDE bond
FREEROTATING
FREEROTATING
PROLINE IMPARTS INFLEXIBILITY ON REGION OF POLYPEPTIDE
The propyl side chain of Proline iscovalently bonded to the beta-nitrogen
to form a ring.
Technically, proline is an IMINO acid
Alpha-carbon at most amino acid residueshave TWO rotatable bonds
Alpha-carbon at proline residueshas only ONE rotatable bond
FREEROTATING
AMINO ACID COMPOSITION AND SOLUTION pH DETERMINE POLYPEPTIDE CHARGE
A protein’sISOELECTRIC POINT (pI)is the pH at which the
protein’s NET CHARGE = 0
An “acidic protein” haspI < 7
A “basic protein” haspI > 7
A protein hasNET NEGATIVE CHARGE
when pH > pI
NET POSITIVE CHARGEwhen pH < pI
pI is determined bya protein’s amino acid
composition.
Eg., an acidic protein has moreacidic residues than basic ones
Different pIs of different polypeptides can be used toseparate these proteins by electrophoresis
at specific pHs.
NEXT LECTURE: PROTEIN STRUCTURE 1Reading: Berg, Chapter 2
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