PROTEIN SYNTHESIS HOW GENES ARE EXPRESSED

BEADLE AND TATUM-1930’S One Gene-One Enzyme Hypothesis

PROTEIN SYNTHESIS GENE (DNA)  TRANSCRIPTION  TRANSLATION  PROTEIN –DNA directs the synthesis of RNA, which is the link between genes and proteins

GENETIC CODE Triplet code –Three nucleotide sequence codes for an amino acid Called a “codon” in mRNA; “anticodon” in tRNA –64 possible combinations code for 20 amino acids There are also “stop” and “start” codes There is redundancy in the code

TRANSCRIPTION Process varies between prokaryotes and eukaryotes –HOW?? Transcription is DNA-directed synthesis of mRNA

TRANSCRIPTION Transcription has 3 stages –Initiation –Elongation –termination

Transcription: Initiation One strand of DNA is used as a template –RNA polymerase: attaches to the promoter site and pulls apart the DNA strands –Transcription occurs in the 5’  3’ direction –Complementary nucleotide bases are added REMEMBER: there is no “T” in RNA; instead, “A” pairs with “U”

TRANSCRIPTION: ELONGATION Nucleotides are added to the 3’ end of the growing mRNA strand About 60 nucleotides/sec are added Length of the primary transcript varies depending on protein being produced

TRANSCRIPTION: ELONGATION

TRANSCRIPTION: TERMINATION RNA polymerase transcribes a terminator sequence in the DNA Details of actual events in termination are still not entirely clear After termination, transcript is called “PRE- mRNA –Modification of mRNA occurs after termination

mRNA Modification 5’ cap is added –Modified guanine nucleotide –Protects the transcript from being broken down by enzymes –Provides a signal for ribosomal attachment Poly-A tail is added to the 3’ end –50 – 250 adenine nucleotides –Inhibits breakdown,assists in attachment, and aids in export out of nucleus

mRNA Modification RNA splicing –Noncoding regions of transcript are removed Called INTRONS –Coding sections are called EXONS

mRNA processing: spliceosomes What makes up a spliceosome?

mRNA processing Why are some parts of mRNA non- coding? In other words, why are there introns? –Perhaps allows flexibility in coding for different proteins, by simply splicing differently Called alternative RNA splicing

TRANSLATION: FROM mRNA to PROTEIN mRNA leaves the nucleus of the cell through nuclear pores Translation occurs in the cytoplasm on a ribosome Translation requires tRNA –Each tRNA has a nucleotide sequence called an ANTICODON that is complementary to the mRNA code –tRNA is made in the nucleus; used repeatedly in the cytoplasm

tRNA

tRNA Each tRNA contains a 3-letter code that codes for an amino acid –What is WOBBLE? Variability in coding of the 3 rd base pairing in the tRNA

TRANSLATION Each tRNA must first pick up its correct amino acid –Joined by aminoacyl-tRNA synthetase –Now called an activated amino acid –Also called “tRNA charging”

TRANSLATION The process occurs at a ribosome Ribosomes-made up of proteins and rRNA

Ribosome P site (peptidyl-tRNA)-holds the tRNA carrying the growing polypeptide chain A site (aminoacyl-tRNA site) holds the tRNA bringing the next amino acid E site (exit site)-where “used” tRNA’s leave the ribosome

TRANSLATION Occurs in 3 stages –Initiation –Elongation –Termination Energy requiring process –Hydrolysis of GTP

TRANSLATION: INITIATION Coming together of mRNA and a tRNA carrying the first amino acid –Complex process –Small ribosomal subunit binds to mRNA and initiator tRNA; then large ribosomal subunit attaches forming the TRANSLATION INITIATION COMPLEX

TRANSLATION: INITIATION

TRANSLATION: ELONGATION Amino acids are added sequentially to the growing polypeptide chain, requiring help of elongation factors Occurs in 3 steps

TRANSLATION: ELONGATION Codon recognition New tRNA is helped into A site Peptide bond formation rRNA (ribozyme) catalyzes a peptide bond forming between the amino acid in the P site and the one in the A site. Growing polypeptide chain is now in the A site Translocation Amino acid in A site is moved to P site tRNA in P site is moved to E site

TRANSLATION: ELONGATION

TRANSLATION: TERMINATION Elongation continues until stop codon in the mRNA reaches the A site UAA, UAG, UGA Release factor binds to the stop codon in the A site –Water molecule is added to the polypeptide chain, thus releasing the polypeptide chain from the ribosome

TRANSLATION: TERMINATION

Polyribosomes Help to speed up process of protein formation, as mRNA can be translated simultaneously by multiple ribosomes

Posttranslational Modification After release from ribosome, polypeptide may require some changes before it is a fully functional protein –Sugars, lipids, phosphates added –Several polypeptides may be joined together

Protein Synthesis: A summary