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Welcome to Bio 28

Introductory Biochemistry

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Clicker Questions1 question per class, 23 classes

1.5 points for participation, 1.5 points for getting itcorrect

Maximum of 60 total points

While the question is on screen (3 min), you maydiscuss your answer with your neighbors.

Only you are allowed to click your clicker, you may

not click another students clicker

You may change your answer up to the time limit

while the question is active

No makeups2

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Brown University Academic CodeExamples of cheating on examinations and quizzes

include, but are not limited to, the following: using

another individual to take an examination in onesplace (taking clicker quiz for another student),bringing into the exam room unauthorized materialsfrom which one gains unfair assistance,

appropriating an exam or exam materials withoutauthorization, missing an exam in order to gain anadvantage, asking for a regrade on an exam thathas been altered after the exam was taken.

All infringements will be sent directly to the Dean ofthe College

Infringement of the academic code entails penalties

ranging from reprimand to suspension, dismissal, or

expulsion from the University. 3

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How are we going to learn biochemistry?

Classes Reading assignments Lehninger 5th edition Weekly conference sessions Sample exams

Getting started Overview of biochemistry Amino acids and peptides

How is this course organized?Canvas course website

Syllabus and calendar Reading Sample exams and answer keys Office hours, TAs, everything else

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Biochemistry

Biochemistry is the chemistry of life.

Biochemistry asks how the remarkable properties of living organisms

arise from the thousands of different biomolecules.

The study of biochemistry shows how the biomolecules that constitute

living organisms interact to maintain and perpetuate life animated solely

by the physical and chemical laws that govern the nonliving universe.

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Properties of living organisms

1. High degree of chemical complexity and microscopic organization.

2. Systems for extracting, transforming, and using energy from the environment.

3. Defined functions for each of an organisms components and regulatedinteractions among them.

4. Mechanisms for sensing and responding to alterations in the surroundings.

5. A capacity for precise self-replication and self-assembly.

6. A capacity to change over time by gradual evolution.

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A high degree of chemical complexityPart 1

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Part 2

A high degree of chemical complexity

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Chemistry of Life

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Only 99% of the mass of most cells

figure 1-12

Essential elements in the chemistry of life

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C is chemically versatile

C can form stable C-C single bonds

C is capable of forming four single bonds

C can form single and double bonds

figure 1-14

Biomolecules are carbon compounds

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Biomolecules are carbon compounds

figure 1-15

R represents any substituent12

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The biochemical ABC

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Macromolecules are biochemical words

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figure 1-11

Structural Hierarchy in the Molecular Organization of Cells

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Where did the biomolecules come from?

1953: The Miller experiment

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The RNA world hypothesis

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Amino acids

Stereoisomers, D- and L-forms; chirality; absolute configuration Optical activity; rotation of the plane of polarized light

General amino acid characteristics, side-chain properties, 3-letter codes,

1-letter codes

Disulfide bonds

UV absorbance of aromatic amino acids; Lambert-Beer law

Zwitterions; pKa; determination of pI

Peptides and peptide bond formation

Levels of protein organization

Partial double-bond character and cis- vs. trans-peptide bonds

Protein sequencing; Edman degradation or Mass Spectrometry

Protein evolution; sequence similarities and phylogenetic trees

Class 1: Outline and Objectives

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L-amino acid

figure 3-2

Amino group

Carboxyl group

Side chain

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Most Amino Acids are Chiral Molecules

figure 1-19

Chiral molecule:Rotated moleculecannot be superimposedon its mirror image

Achiral molecule:Rotated moleculecan be superimposedon its mirror image

greek chiros = 20

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D- and L-Enantiomers

figure 3-321

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Absolute Configuration

figure 3-4Glyceraldehyde is the standard22

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R, S nomenclature

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There are 20 amino acids that normally occur in proteins.

All chiral amino acids that occur in proteins have the L

configuration.

figure 3-2

These 20 amino acids can be grouped according to thechemical nature of their side chains (R groups).

Amino group

Carboxyl group

Side chain

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figure 3-5

Ala AGly G Val V

Ile I Met MLeu L

Pro P

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Ala AGly G Val V

Ile I Met MLeu L

Pro P

Note: Glycine is not chiral

figure 3-526

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Note: Isoleucine has two chiral centers

(the L-isomer with the opposite configuration at C3 is called L-alloisoleucine)

figure 3-5

Ala AGly G Val V

Ile I Met MLeu L

Pro P

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figure 3-5

Asp D Glu E

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figure 3-5

Histidine is conditionally positively charged

H+

His HArg RLys K

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(which one of these has a possible alloform?)figure 3-5

Ser S Thr T Cys C

Asn N Gln Q

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Oxidation of Cysteine Results inDisulfide Bond Formation

figure 3-7

Cysteine

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figure 3-5

Phe F Tyr Y Trp W

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figure 3-6

Absorbance of UV Light by Aromatic Amino Acids

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C

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Absorbance and Absorption CoefficientsLambert-Beer Law

A(bsorbance) = log = log 1/TI

I

0

Transmission =I

I0

100% T 0%

box 3-1

Example of the relationship of T to A:

If 90% of the incident light is absorbed after 1 cm, log[100/10] = 1.0 = A;

if 99% is absorbed, log[100/1] = 2.0 = A.34

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Lambert-Beer Law

A is linearly proportional to cand l.

is the molar absorption [or extinction] coefficient (units are M1 cm1).

Application: Determination of the concentration of a pure protein:

c= A / l

A is measured (typically at 280 nm);

lis constant (1 cm)

280 is known ( of the protein is the sum of the s of all Trp and Tyr residues).

Assume that the protein has an average number of Trp and Tyr residues

compared to all known protein sequences

A(bsorbance) = log = log 1/T = clI

I

00 A

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Nonionic and Zwitterionic Forms

figure 3-9

A zwitterion can act as either anacid or a base.

Substances that have this

property are amphoteric and are

often called ampholytes

(amphoteric electrolytes). Free amino acids are zwitterionic

at neutral pH.

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pH = pK + log[A]

[HA]

Amino Acid Titration,

Ionization Statesand Isoelectric Point (IEP)

figure 3-10

pI = (pK + pK )1

221

a

pI: The pH at which the net

electric charge of a molecule is

zero.

+1 0 1 net charge

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figure 3-12a

+1 0 1 2 net charge

Titration Curvefor Glutamate

pI = (4.25 + 2.19) = 3.221

2

pI

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Peptide Bond Formation

figure 3-13

Condensation reactionHydrolysis

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Oligopeptides

figure 3-15

This is: Ala-Glu-Gly-Lys (orAEGK)

Not: Lys-Gly-Glu-Ala (orKGEA)

Always written beginning atthe end with the free NH3

+(the N-terminal end)

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Partial Double Bond Character of Peptide Bonds

figure 4-2a

.

The partial double-bond character makes the peptide bond planar

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Hierarchy of Protein Organization

figure 3-1642

P i f i

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Primary structure of proteins:protein sequencing

figure 3-15, 3-2443

S h d d Ed d d i

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Sangers method and Edman degradation

figure 3-2544

S i

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Sequencinglarger proteins

figure 3-2745

P tid i b t d t t

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Peptide sequencing by tandem mass spectrometry

box 3-246

Sequencing with MS/MS

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Sequencing with MS/MS

S H N S A L pY S Q V Q K88 225 339 426 497 610 853 1068 1167 1299 1459940

1459 1372 1234 1121 1033 962 849 606 519 391 292 164

b ion:

:y iony1y12

b1 b12

1) Each peptide collisional-induced dissociation produces only two fragments2) Cleavage can occur at any peptide backbone amide bond

box 3-247

P t i li t

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Protein sequence alignment

Consensus sequences reflect the mostcommon amino acid at each position.

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E l ti t b d l i

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Evolutionary trees based on sequence analysisof protein families

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A consensus tree of life

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A consensus tree of life

Comparison of many protein sequences between50