* 17 Flashcards
Beadle and Tatum’s experiment
- bombarded bread mold Neurospora crassa w/ X-rays to cause genetic changes, then looked among the survivors for mutants that differed in their nutritional needs from the wild type
- wild type can grow in the lab on a simple sol’n of inorganic salts, glucose, and the vitamin biotin, incorporated into AGAR, a support medium (minimal medium)
- nutritional mutants were grown on a complete growth medium, which consisted of minimal medium + all 20 amino acids + other nutrients
codon start/stop
UAA stop
UGA stop
UAG stop
AUG start / methionine
reading frame
On an mRNA, the triplet grouping of ribonucleotides used by the translation machinery during polypeptide synthesis.
promoter
- the DNA sequence where RNA polymerase attaches and initiates transcription
- includes within it the transcription START POINT, the nucleotide where RNA synthesis actually begins
- typically extends several dozen or more nucleotide pairs upstream from the start point
- typically includes a crucial sequence called a TATA box (TATA is the sequence on the nontemplate strand)
transcription unit
A region of DNA that is transcribed into an RNA molecule
transcription initiation complex
- a collection of proteins called TRANSCRIPTION FACTORS mediate the binding of RNA polymerase and the initiation of transcription; only after transcription factors are attached to the promoter does RNA pol II bind to it
- the whole complex of transcription factors + RNA pol II
RNA polymerase and types
- pries the two strands of DNA apart and joins together RNA nucleotides complementary to the DNA template strand
- unlike DNA pols, RNA pols are able to start a chain from scratch, no primer needed
- binds in a precise location and orientation on the promoter, therefore determining where transcription starts and which of the two strands of the DNA helix is used as the template
- bacteria: a single type of RNA pol that synthesizes all RNA types
- eukaryotes: at least 3 types; the one used for mRNA synthesis is called RNA pol II
transcription elongation
- RNA pol exposes about 10 to 20 DNA nucleotides at a time for pairing w/ RNA nucleotides
- rate: 40 nucleotides per second in eukaryotes
transcription termination in bacteria
- transcription proceeds thru a terminator sequence in the DNA
- the transcribed terminator (an RNA sequence) functions as the termination signal, causing the polymerase to detach from the DNA and release the transcript
- the transcript requires no further modification before translation
transcription termination in eukaryotes
- RNA pol II transcribes a sequence on the DNA called the POLYADENYLATION SIGNAL sequence, which codes for a polyadenylation signal (AAUAAA) in the pre-mRNA
- this is also the 3’ UTR
- then, at a point 10 to 35 nucleotides downstream from the AAUAAA signal, proteins associated w/ the growing RNA transcript cut it free from the polymerase, releasing the pre-mRNA
- the pre-mRNA then undergoes processing
RNA processing
- in eukaryotes, the modification of pre-mRNA before the mRNA is dispatched to the cytoplasm
- both ends of the primary transcript are altered
- certain interior sections of the RNA molecule are cut out and remaining parts spliced together
alteration of mRNA ends
- 5’ end is synthesized first; it receives a 5’ cap, a modified form of a guanine nucleotide added onto the 5’ end after transcription of the
- at the 3’ end, an enzyme adds 50 to 250 more adenine nucleotides, forming a POLY-A TAIL
- 3’ and 5’ UTRs (untranslated regions) are on the interior side of the altered ends; the 3’ UTR is the polyadenylation signal
function of altered mRNA ends
- facilitate the export of the mature mRNA from the nucleus
- help protect mRNA from degradation by hydrolytic enzymes
- help ribosomes attach to the 5’ end once the mRNA reaches the cytoplasm
RNA splicing numbers
- avg length of a transcription unit along a human DNA molecule is about 27,000 np
- however, it only takes 1,200 nucleotides in RNA to code for the avg-sized protein of 400 AA
introns
intervening sequences; noncoding segments of nucleic acid that lie btwn coding regions
exons
- eventually expressed, usually by being translated into AA sequences
- exceptions include the UTRs of the exons at the ends of the RNA, which make up part of the mRNA but aren’t translated into protein
- exons are sequences of RNA that exit the n ucleus
snRNPs
- particles called small nuclear ribonucleoproteins that recognize the signal for RNA splicing, a short nucleotide sequence at each end of an intron
- located in the nucleus; RNA + protein molecules
- the RNA particle is called a small nuclear RNA, about 150 nucleotides long
spliceosome
- several diff snRNPs + additional proteins
- almost as big as a ribosome
- interacts w/ certain sites along an intron, releasing the intron, which is rapidly degraded, and joining together the 2 exons that flanked the intron
- snRNAs catalyze these processes, as well as participating in spliceosome assembly and splice site recognition
ribozymes
- RNA molecules that function as enzymes
they’re able to do so b/c - b/c RNA is single-stranded, a region of an RNA molecule may base-pair w/ a complementary region elsewhere in the same molecule, which gives the molecule a particular 3D structure
- like certain AA in an enzymatic protein, some of the bases in an RNA contain functional groups that may participate in catalysis
- the ability of RNA to H-bond w/ other nucleic acid molecules adds specificity to its catalytic activity
alternative RNA splicing
- many genes are known to give rise to 2 or more diff polypeptides, depending on which segments are treated as exons during RNA processing
- b/c of this, the number of diff protein products an organism produces can be much greater than its number of genes
domains
- proteins often have a modular architecture consisting of discrete structural and functional regions called domains
- 1 domain might include the active site, while another might allow the enzyme to bind to a cellular membrane
- in quite a few cases, diff exons code for the diff domains of a protein
exon shuffling
- introns increase the probability of crossing over btwn the exons of alleles of a gene, simply by providing more terrain for crossovers w/o interrupting coding sequences
- while most changes would be nonbeneficial, occasionally a beneficial variant might arise
tRNA molecule
- single RNA strand only about 80 nucleotides long (compared to the hundreds of nucleotides for most mRNA molecules)
- b/c the presence of complementary stretches of nucleotide bases that can hydrogen-bond to each other, this single strand can fold back upon itself and form a molecule w/ a 3D structure
- 2D: looks like cloverleaf – 4 base-paired regions, 3 of which are loops
- 3D: roughly L shaped
- loop at one end of the L: anticodon
- other end (just a strand, NOT looped): 3’ end, attachment site for amino acid
first instance of molecular recognition
- matching up of tRNA and amino acid
- aminioacyl-tRNA synthetases, a family of related enzymes; 20, one for each AA
- their active sites fit only a specific combination of amino acid and tRNA
1. synthetase’s active site binds the amino acid and ATP
2. ATP loses 2 phosphate groups and bonds to the AA as AMP
3. appropriate tRNA covalently bonds to amino acid, displacing AMP
4. the resulting aminoacyl tRNA, charged w/ AA, is released by the enzyme