protein metabolism l.
Download
Skip this Video
Loading SlideShow in 5 Seconds..
Protein Metabolism PowerPoint Presentation
Download Presentation
Protein Metabolism

Loading in 2 Seconds...

play fullscreen
1 / 87

Protein Metabolism - PowerPoint PPT Presentation


  • 830 Views
  • Uploaded on

Protein Metabolism. The genetic code Protein synthesis (translation) Protein targeting & degradation. What are needed for protein synthesis? >70 ribosomal proteins >20 enzymes to activate amino acid precursors >12 proteins/enzymes for the initiation, elongation & termination

loader
I am the owner, or an agent authorized to act on behalf of the owner, of the copyrighted work described.
capcha
Download Presentation

PowerPoint Slideshow about 'Protein Metabolism' - menefer


An Image/Link below is provided (as is) to download presentation

Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author.While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server.


- - - - - - - - - - - - - - - - - - - - - - - - - - E N D - - - - - - - - - - - - - - - - - - - - - - - - - -
Presentation Transcript
protein metabolism
Protein Metabolism
  • The genetic code
  • Protein synthesis (translation)
  • Protein targeting & degradation
slide2

What are needed for protein synthesis?

>70 ribosomal proteins

>20 enzymes to activate amino acid precursors

>12 proteins/enzymes for the initiation, elongation & termination

of polypeptides

~100 enzymes for the final processing of proteins

>40 tRNAs & rRNAs

The most complex biosynthetic process!

slide3

Three major advances for the understanding

of protein synthesis

1) Paul Zamecnik et al., 1950s (p.1021)

newly synthesized, radiolabeled proteins are accumulated

at small ribonucleoprotein particle (i.e., ribosome) in liver.

slide4

Three major advances for the understanding

of protein synthesis (cont’d)

2) Mahlon Hoagland & Zamecnik

activated amino acids are attached to a heat-stable soluble RNA

(i.e., tRNA), forming aminoacyl-tRNAs.

Aminoacyl-tRNA synthetases are involved.

slide5

3) Francis Crick’s adaptor hypothesis

a small nucleic acid (perhaps RNA) could serve the role of an

adaptor, one part binding a specific a.a. and aother part recognizing

the nt sequence encoding that a.a. in the mRNA.

slide6

The triplet, nonoverlapping code

Insertion or deletion mutations alter the sequence of triplets.

Adding/subtracting 3 nt leaves the remaining triplet intact,

providing evidence that a codon has 3 nt.

slide7

codon: a triplet of nucleotides that codes for a specific amino acid.

reading frame: established by the first codon, then begins a new one

every 3 nucleotide residues.

slide8

The Genetic Code Was Cracked

Using Artificial mRNA Templates

Marshall Nirenberg, 1961

enzymatic methods to synthesize

poly(U) > phenylalanine

poly(C) > proline

poly(A) > lysine …

Nirenberg & Philip Leder, 1964

trinucleotides induce specific binding of aminoacyl-tRNA

to ribosome.

H. Gobind Khorana, 1960s

chemical methods to synthesize polynucleotides with

repeating sequences of 3 & 4 bases > polypeptides

slide9

“Dictionary” of amino acid code words

as they occur in mRNAs.

termination codon (red)

initiation codon (green)

slide10

Each reading frame gives a different sequence of codons,

but only one is likely to encode a given protein.

Open reading frame (ORF):

a reading frame without a termination codon among

50 or more codons.

slide11

Codon is degenerate:

an amino acid

may be specified

by more than one codon.

slide13

Codon pairing relationships when the tRNA

anticodon contains inosinate

protein metabolism14
Protein Metabolism
  • The genetic code
  • Protein synthesis (translation)
  • Protein targeting & degradation
slide17

“Dictionary” of amino acid code words

as they occur in mRNAs.

termination codon (red)

initiation codon (green)

slide18

EF-Tu

EF-G

GDP (red)

C-terminus (green)

mimics tRNA

tRNA (green)

slide19

Overlapping Genes in Different Reading Frames

Are Found in Some Viral DNAs

fX174

slide20

Overlapping Genes in Different Reading Frames

Are Found in Some Viral DNAs:

Genes within genes

slide22

The Ribosome Is a Complex

Supramolecular Machine

Masayasu Nomura et al., 1960s (p.1037)

both ribosomal subunits can be broken down into their

RNA and protein components, then reconstituted in vitro.

slide23

Structure of the bacterial ribosome

at near-molecular resolution

slide25

Ribosomal subunits are identified by their S (Svedberg unit)

values, sedimentation coefficients that refer to their rate of

sedimentation in a centrifuge.

slide29

The sequences of the rRNAs of many organisms have been

determined. Each has a specific three-dimensional

conformation featuring extensive intrachain base pairing.

Models for the secondary structure

of E. coli 16S and 5S rRNAs

slide30

Transfer RNAs Have Characteristic Structure Features

Robert H. Holley et al., 1965 (p.1038)

yeast tRNAAla

cloverleaf conformation 苜蓿葉形

slide32

Three-dimensional structure of yeast tRNAPhe

deduced from X-ray diffraction analysis

protein synthesis
Protein Synthesis
  • Stage1:Aminoacyl-tRNA synthetases attach the correct amino acids to their tRNAs
  • Stage 2: A specific amino acid initiates protein synthesis
  • Stage 3: Peptide bonds are formed in the elongation stage
slide36

Amino acid + tRNA + ATP

Mg2+

Aminoacyl-tRNA synthetase

aminoacyl-tRNA + AMP + PPi

slide37

Proofreading by aminoacyl-tRNA synthetase

e.g., Ile-tRNAIle synthetase favors activation of Ile over Val

by a factor of 200, i.e., it distinguishes between Val and Ile.

slide38

Interaction between an aminoacyl-tRNA synthetase

and a tRNA: a “second genetic code”

Recognition sites by:

unique enzyme (orange)

several enzymes (green)

all enzymes (blue)

slide39

Gln-tRNA synthetase

Asp-tRNA synthetase (dimeric)

tRNA (green)

bound ATP (red)

slide40

The tRNAAla elements recognized by the

Ala-tRNA synthetase are usually simple.

Just a single G=U base pair (red)!

Synthetic simple

form also works!

protein synthesis41
Protein Synthesis
  • Stage1: Aminoacyl-tRNA synthetases attach the correct amino acids to their tRNAs
  • Stage 2:A specific amino acid initiates protein synthesis
  • Stage 3: Peptide bonds are formed in the elongation stage
slide42

Howard Dintzis, 1961

polypeptides grow by addition of new amino acid

to the carboxyl end

slide43

synthetase

Met + tRNAfMet + ATP

Met-tRNAfMet + AMP + PPi

N10-Formyltetrahydrofolate + met-tRNAfMet

tetrahydrofolate + fMet-tRNAfMet

transformylase

tRNA

The distinction between

initiating AUG and internal one

is straightforward...

slide44

Three steps of initiation:

Aminoacyl site

Peptidyl site

Initiation Factor (IF)

the initiation complex forms in

the expense of the hydolysis of

GTP to form GDP and Pi.

slide45

The initiating AUG is guided by

the Shine-Dalgarno sequence

in the mRNA

slide46

Protein complexes in the formation of

a eukaryotic intiation complex

eIF

protein synthesis48
Protein Synthesis
  • Stage1: Aminoacyl-tRNA synthetases attach the correct amino acids to their tRNAs
  • Stage 2: A specific amino acid initiates protein synthesis
  • Stage 3:Peptide bonds are formed in the elongation stage
slide49

Peptide Bonds Are Formed

in the Elongation Stage

  • Elongation requires:
  • the initiation complex
  • aminoacyl-tRNAs
  • elongation factor (EF-Tu, -Ts,-G)
  • GTP

Proofreading on the ribosome:

EF-Tu.GTP/EF-Tu.GDP complexes

(~milliseconds) provide opportunities

for the codon-anticodon interactions.

Elongation step 1:

Binding of the second aminoacyl-tRNA

slide50

Elongation step 2:

Formation of the first

peptide bond

Peptide transferase

slide51

Elongation step 3:

Translocation

slide53

How to know GTP is involved?

GTP analog

slows hydrolysis, improving the fidelity (by increasing

the proofreading intervals) but reducing the rate

of protein synthesis.

slide54

Termination of polypeptide synthesis

requires a special signal

Release (or termination) factor

slide56

Energy cost of fidelity in protein synthesis

More than 4 high-energy bonds are required for

the formation of each peptide bond of a polypeptide:

2 ATP/GTP during aminoacyl-tRNA formation

2 GTP during the first elongation step & translocation

slide57

Rapid translation of a single mRNA

by polysome in both prok./euk. cells

Polysome:

a fiber between adjacent ribosomes

in the cluster of 10 to 100.

slide60

Posttranslational modification

e.g., modification of individual amino acids:

phosphorylation

slide63

Posttranslational modification (cont’d)

e.g., addition of isoprenyl groups

slide64

Protein Synthesis Is Inhibited by

Many Antibiotics and Toxins

e.g., disruption of peptide bond

formation by puromycin

Peptidyl puromycin

protein metabolism66
Protein Metabolism
  • The genetic code
  • Protein synthesis (translation)
  • Protein targeting & degradation
slide67

George Palade...

David Sabatini & Gunter Blobel, 1970

Signal sequences of some eukaryotic proteins:

Signal recognition particle (SRP)

slide68

Directing eukaryotic proteins with the appropriate

signals to the endoplasmic reticulum (ER)

slide70

Glycosylation Plays a Key Role in Protein Targeting

Synthesis of the core oligosaccharide of glycoproteins

slide72

Tunicamycin mimics the structure of

UDP-N-acetylglucosamine (UDP-GlcNAc) and blocks

the first step of glycosylation.

slide76

Proteins Are Targeted to Mitochondria and

Chloroplasts by Similar Pathways

slide78

Signal Sequences for

Nuclear Transport

Are Not Cleaved

slide82

Cells Import Proteins by Receptor-Mediated Endocytosis

clathrin

coated pit

Joe Goldstein

& Mike Brown

slide84

Protein Degradation Is Mediated by

Specialized Systems in All Cells

Proteasome: (Mr 1 x 106)

The ATP-dependent, ubiquitin

involved proteolytic system

in eukaryotes.

Three-step cascade pathway

by which ubiquitin is attached to

a protein.

slide87

The Nobel Prize in Chemistry 2004

"for the discovery of ubiquitin-mediated protein degradation"

Aaron

Ciechanove

Avram

Hershko

Irwin Rose