Protein and Protein Synthesis

Question 1

what are proteins

Answer 1

-polypeptides
-comprised of polymers of AA connected by peptide bonds
-peptide bonds are between carboxylic acid of one amino acid and the amino group of the next amino acid
-proteins are directional running from N terminus to C terminus

Question 2

AA structure

Answer 2

-have an amino grpup on one side, alpha carbon in the middle and a carboxyl group on the other side

Question 3

the 20 AA

Answer 3

-there are 20 different AA that make up proteins that have the same backbone but different R groups
-they can be grouped by the properties of their R groups such as non-polar, polar, positively charges and negatively charged

Question 4

protein structure

Answer 4

-primary structure = AA sequence
-secondary structure - alpha helices or beta sheets formed by hydrogen bonding in peptide backbne
-tertiary structure - 3D structure, includes multiple secondary strucutral elements arranged in different ways and other structural features
-quaternary structure - result of multiple different polypeptides coming together into multimeric proteins

Question 5

diagram of alpha helices, beta sheets

Answer 5

Question 5

tRNA

Answer 5

-ribosome uses tRNAs to convert the mRNA sequence into a proteins equence
-around 70-95 bases long
-extensively strucutreed
-often contain modifeid bases
-each tRNA has a specific anticodon that bind a particular three base codon
-at the other end, tRNAs carry the specific AA that corresponds to that codon

Question 6

tRNA synthases

Answer 6

-enzymes that charge tRNAs
–add the AA to the CCA at the 3’ end

Question 7

the genetic code

Answer 7

-each codon corresponds to a specific AA or to a stop codon
-there are 64 possible codons and 20 AA
-multiple different codons can encode a paritucular AA
-in other cases certain tRNAs can work even if there os a songle mismatch in 3rd position (wobble) - same tRNA for 2 different codons

Question 8

start codon

Answer 8

-encodes first Amino acid of a aparticular ORF
-where translation begins
-it is typically an AUG for all three domains of life
-alternative start codons are GUG and UUG
-when AUG is encountered during normal translation it encodes methionine
-used in archaea and eukary unmodified

Question 9

start codon of bacteria

Answer 9

-N-formylmethionine (fMet)
-chemically modified version of methione using special tRNA

Question 10

the ribosome

Answer 10

-prokaryotic - 70S - made up of 2 subunits 30S subunit and 50S subunit
-each subunit is comprised of rRNA and ribosomal proteins
-t RNAs and several other proteins interact with ribosome transiently - perform important roles in translation
-rRNA carries out much of the main function of the ribosome including catalyzing peptide bone formation

Question 11

E.coli ribosome

Answer 11

30S ribosome contains 16S rRNA and 21 proteins
-50 S contains 5S/23S rRNA and 31 peoteins

Question 12

initiation of translation

Answer 12

-RNA has many AUGs
-to locate bona fide start codons bacterial mRNAs use a ribosome binding site
-RBS is an RNA sequence that is recognized by ribosome - must be appropriately spaced from a start codon
-16D rRNA component of a free 30S ribsomal subunit binds to RBS to initiate translation

Question 13

3 tRNA binding sites of the ribosome for translation

Answer 13

-A site- where new charges tRNAs enter and recognize the codon being translated. Once in place the growing peptide from P site is transferred its AA and a peptide bond forms
-P site - after bond formation, translocation occrs, RNA moves 3 bases (one codon). The tRNA from A site moves to P site. This tRNA transfers into growing AA to the new charged tRNA that has entered the A site
E site - uncharged tRNAs exit here

Question 14

what is the energy source for translation

Answer 14

-GTP

Question 15

diagram of translation

Answer 15

Question 16

terminating translation

Answer 16

hits stop codons UAA, UGA, or UUAG
-once this occurs, a protein called a release factor bind
-this results in ribsome releasing the mRNA ans the completed polypeptide
-30S/50S subunits disscoiate. Ribsoosme is free to begin new round of translation

Question 17

polysomes

Answer 17

-multiple ribsomes on a sinle transcript
-same mRNA can be simultaenously translated by multiple different ribsoosmes
-as 5’ end of RNA in an ORF is translated and freed up a new ribsome can bind and begin making protein before previous ribsome is finished, Each ribosome on a polysome builds a complete protein

Question 18

coupling of transcription and translation

Answer 18

in prokaryotes transcription and translation are often coupled
-RNA being translated while transcription is still going on

Question 19

eukaryotic translation

Answer 19

-overall very similar to prokaryotes
in eukaryotes transliation and transcription are not coupled and occur in different compartments (transcr in nucleus transl in cytoplasm)
-each eukaryotic mRNA encoes one gene, many prokaryotic transcripts enncode 2+ ORF
-main mechanistic difference is the initaiton step. 5’ recognized in prokaryotes
-eukaryotic ribsomes are larger and a bit more complex - 40S and 60S, 80S in complete ribosome

Question 20

chaperones

Answer 20

-some proteins require chaperones to fold into correct conformation
-others can dols LONE IN MOST SITUATIONS BUT ANY PROTEIN CAN MISFOLE
-CHAPERSONES ARE PROTEINS THAT HELP OTHER PROTEINS ADOPT THEIR PROPERLY FOLDED AND FULLY ACTIVE STATE
-ALL 3 DOMAINS REQUIRE CHAPERNES
-UBUTUAK FIKDUBG, REFOLDUGB DEBATURED PROTEINS, HELPING SUBUNITS IN MULTIMERIC PROTEINS COME TOGETHER
-THERE ARE RNA CHAPERONES THAT HELP RAS ADOPT THE CORREXT STRUCTURE
-SPECIAL CHAPERones are activated in response to high or low temps to assist with protein and RNA folding

Question 21

Ecoli chaperones

Answer 21

-Dnah/DnaK and GroEL/GroES are major chaperones tat assist in protein folding- essential for survival
-among most abundant proteins in cell
-ATP hydrolysis for energy
-enable unfolded or unstable conformations to refold in controlled fashion

Question 22

protein secretion in prokaryotes

Answer 22

-all proteins are syntheiszed by pribsomez in the cytoplasm
-but many preotiins are required in other locations
-use translocase systems that transport proteins across and into the cytoplasmic membrane

Question 23

translocated proteins

Answer 23

• most translocated proteins contain a signal sequence at the N terminud that targets the protein to a parituclar secretion system - often removed after translocation
  -the sec secretion system and twin arginine translocase are uiquotous in prokaryotes- others are more specialized
  -eukaryotic protein folding, processing/modification and transport systems are complex and not covered in this course

Question 24

the Sec system

Answer 24

-Tat pathway secretes prefoled proteins across membrane
-the sec secretion system recognizes a signal sequene in rhw first amino CIDS OF A PROTEIN - TRANSlocated unfolded protein before it folds

Question 25

in the sec pathway what are the two paths proteins can take

Answer 25

-passed across cytoplasmic membrane (SecA pathway)
-recognized by RNA/protein complex-signal recognition particle - and inserted into the cytoplasmic membrane (SRP pathway)

Question 26

similarities between SRP and SecA pathway

Answer 26

-pass the unfolded protein through membrane channel (YEG translocon)
-require ATP dor energy
-signal sequnce is cleaved following translocation

Question 27

the sec system diagram

Answer 27