Chapter 12 From DNA to Protein: Genotype to Phenotype

Biology 101Tri-County Technical CollegePendleton, SC

From past to present…

One gene = one proteinOne gene = one enzyme Beadle and Tatum Altered phenotype = altered enzyme

protein

One gene = one polypeptide**Function of one gene is to control (direct) the synthesis (production) of ONE polypeptide

Comparing RNA to DNA

RNA usually consists of only ONE polynucleotide strand Chargaff’s rule NOT applicablePentose is riboseAUGCAlways constructed as single strand but may fold over and base pair with itself

Central Dogma

States DNA codes for production of RNA, RNA codes for production of protein, and protein does NOT code for production of protein, RNA, or DNATook a long time to get to here…remember Hershey and Chase’s bacteriophage experiments

Central Dogma Visual

Making Stuff…so to speak

RNA molecule forms as complementary copy of one DNA strand of a particular gene in process called transcriptionForms messenger RNA (mRNA) which travels to cytoplasm where it serves as template for protein synthesisAdapter molecule (Crick) that can bind a specific AA and recognize sequence of nucleotides with another region is called transfer RNA (tRNA) Sequence of 3 bases called ANTICODON

Stuff, cont.

tRNA adapter line up on mRNA so that AA are in proper sequence for growing polypeptide chain in process called translationTranslation requires third type of RNA called ribosomal RNA (rRNA) which is RNA complexed with proteinsGiven gene (sense) transcribed into mRNA; tRNAs translate sequence of bases in mRNA into appropriate sequence of AAs in assocation with ribosomes (rRNA and protein)

Making Stuff Visual

3 Steps of Transcription

Initiation, elongation, and terminationBegins at promoter (sequence of DNA to which RNA polymerase binds At least one promoter for each gene

Part of promoter is initiation site where transcription beginsRNA polymerase moves in 3’ to 5’ directionWhich means that transcription proceeds in 5’ to 3’ direction ONLY

Steps, cont.

RNA polymerase unwinds DNA about 20 bases at a timeRNA polymerase then adds RNA nucleotides to the 3’ end of growing strand of RNATermination site reached, RNA polymerase detaches from DNAUnlike DNA polymerases, RNA polymerases DO NOT inspect their work

Steps, cont.

Transcription errors = one mistake for every 103 to 105 basesChalk talk time on transcriptionOne can bet the farm an opportunity will be provided to demonstrate understanding of this process

Understanding the codon

Codon is sequence of 3 bases on mRNA that “code” (specify) a particular AA or a stopThe genetic code will be providedAUG (methionine) is universal start codonUAA, UAG, and UGA are stop codonsMUST know how to read/apply the codeCare to guess how they figured out how many bases must be in a codon?

I like the term degenerate…

There are 64 different 3 letter codons61 of them call for a specific AAAn AA may be represented by more than one codon, so we now say code is redundantUnambiguous means that specific codon requires a specific AAIt is NOT roll your own, or any ole AA will doMore than one way to say “put valine here” but NOT any ole AA here

Putting it together

Given a sequence bases in DNA (either sense or nonsense) build the Doc everything else.One can bet the farm this will show up again (and again)Hot damn, it be chalk talk time again

Ribosome structure and function

Each ribosome consists of 2 subunits: small and largeDifferent proteins and RNA’s in subunit held together by ionic and hydrophobic forcesRibosome can combine with any mRNA and all tRNAs and can be used to make different polypeptide products

Ribosomes, cont.

Ribosome simply molecular factory where proteins are assembledLarge subunit contains 4 sitesT (transfer) site is where tRNA carrying an amino acid first lands on ribosomeA (amino acid) site is where tRNA anticodon binds to mRNA codon, lining up correct AA to be added to growing chain

Ribosomes, cont.

P (polypeptide) site is where tRNA adds its amino acid to growing chainE (exit) site is where tRNA, having given up its AA resides before leaving ribosome**Codon-anticodon interactions and peptide bond formation occur ONLY at A and P sitesPolysome: more than one ribosome translating a strand of mRNA70S and 80S ribosomes and endosymbiotic theory of eukaryotic evolution

Take three steps and translate

Initiation, elongation, and terminationInitiation depends on initiation factors (proteins) which direct process and use GTPEnergy for elongation also comes from GTPTermination depends on “stop” codonStop codon does NOT bind any tRNA but does bind release factor which causes water molecule to be added to PP chain instead of AACauses separation of complete protein from ribosomeHot damn…chalk talk time on translation

Protein Destination

All protein synthesis begins on free ribosomes in cytoplasmAs PP chain made, into in its AA sequence gives it one of two sets of directions Finish translation and be released into cytoplasm Stop translation and go to rough ER and finish

there

Can be sent anywhere in cell, or lacking specific instructions, it is secreted from cellAfter translation, some PPs have short exposed sequence of AAs that act like postal “zip code”

Destination, cont.

Signal sequences are at N terminus or in interior of AA chain Have conformation that allow binding to

specific receptor proteins (docking proteins) on outer membrane of appropriate organelle

Receptor forms channel in membrane

If specific hydrophobic sequence of ~25 AAs occurs at beginning of PP chain, finished product destined for ER, lysosomes, PM, or secretion from cell

Destinations, cont.

Sugars can be added as part of addressAddressing of protein to destination is property of its AA sequence and so is genetically determinedMucoplidosis II (I-cell disease) caused by lack of essential enzyme for formation of lysosomal targeting signal

Destinations, cont.

Proteins destined for lysosome never get there but stay in Golgi apparatus where they form I (inclusion) bodies or are secreted from cellInability to perform normal lysosome functions leads to progressive illness and death in childhood

Protein Modification

**Review of precursor conceptMost proteins modified both covalently and noncovalently Essential for final functioning of protein

Proteolysis is cutting of PP chain Some proteins made from polyproteins

cut into final products by enzymes called proteases

Modification, cont.

Glycosylation is adding of sugar to protein Important in addressing; 3-dimensional

structure; recognition of proteins at cell surface; and in stabilizing stored proteins

Phosphorylation is addition of phosphate group to protein Change 3-dimensional structure (often

exposing active site or binding site for another protein

Making sense of nonsense…

Mutations are heritable changes in genetic information True, but…sudden, permanent

change in DNA

Point mutations = mutations of single genesOne allele becomes another because of small alterations in sequence or number of nucleotides

Making sense, cont.

Point mutation result from addition or subtraction of a base or the substitution of one base for another in DNA (exon/intron)Genetic code is redundant so some point mutations result in NO change in AA sequence and are called silent/synonymousSome base substitution mutations change message so one AA substituted for another and are called missense

Making sense, cont.

Sickle-cell disease results from ONE wrong AA Missense may cause protein NOT to function but usually just reduce efficiencyNonsense mutation cause a “STOP” codon to be inserted in mRNA to be translatedResults in nonfunctional productFrame shift mutation occurs when base pairs are inserted into or deleted from DNAThrow message out of synchShift the reading frame-bad news, big time

Some Key Terms

One more time…with feeling…yehaw!!Chromosomal mutations change position or direction of DNA segment without removing any genetic info or may cause segment to be irretrievably lostLet’s talk karyotype**Review: deletion, duplication, inversion, and translocation