Exam "4": Translation, Control of Gene Expression, Population Genetics (Bio 375 - Genetics) Flashcards
amino acid components
carboxyl group, amino group, and R group
identity of amino acid is determined by
the R group attached to the alpha carbon
in H2O (physiological conditions), the true form of amino acid is
charged, with the amino group bearing a positive charge (+NH3) and the carboxyl group bearing a negative charge (COO-)
primary protein structure
sequence of amino acids
secondary protein structure
folded/twisted polypeptide chain (beta pleated sheet and alpha helix)
tertiary protein structure
secondary structure folded further
quaternary protein structure
2+ polypeptide chains associated
polypeptides
amino acids are joined to each other via peptide bonds formed via dehydration synthesis
dehydration synthesis
combining two compounds through removal of water molecules– forms peptide bonds between amino acids, phosphodiester bonds between nucleic acids
hydrolysis
breaking of bonds through addition of water
tRNA
serves as translating molecule between nucleic acids and amino acids… has characteristic structure with an anticodon and amino acid attachment site
codon
found in the mRNA, combinations of three nucleotides – there are 61 “sense” trinucleotide sequences that code for amino acids and 3 “nonsense” trinucleotide sequences that are stop codons
ribosomes read nucleotides of mRNA
in groups of three (codons) from 5’ -> 3’
AUG- Met
“start” codon when at the beginning of mRNA, but when later in mRNA sequence codes for the amino acid Methionine
sense codons
61 codons that code for amino acids
nonsense codons
3 stop codons (UAA, UAG, UGA)
degenerate code
amino acids may be specified by more than one codon
synonymous codons
codons that specify the same amino acid
isoaccepting tRNAs
tRNAs that accept the same amino acid but have different anticodons
wobble position
third position of codon base pairs weakly with anticodon, where nonconventional base pairing is permitted…. allows the last position of the anticodon to pair weakly with the codon
tRNAs can recognize
1-3 different codons
I (inosine)
a nitrogenous base only found in tRNA; can base pair with A, U, or C (due to wobble position)
reading frame
way in which the codon sequence of a mRNA is read… codons are nonoverlapping so each nucleotide is part of a single codon… is set by position of initiation codon
initiation codon
first codon of mRNA to specify an amino acid… usually the first AUG in mRNA – which is N-formylmethionine (modified methionine inserted on “start” AUG) and is removed post translationally
prokaryotic translation
takes place in ribosome… ribosome attaches to 5’ end of mRNA and proceeds 3’ down the chain… polypeptide is synthesized starting at amino end (N -> C), with elongation continuing by adding amino acids to carboxyl end
translation stages
- tRNA loading/charging… 2. initiation… 3. elongation… 4. termination… [with steps 2-4 involving the ribosome]
translation step 1. tRNA loading
tRNA molecules must be loaded with correct amino acid (each tRNA is specific for a certain amino acid)– amino acids are loaded onto 3’ end of the tRNA molecule as specified by enzyme aminoacyl-tRNA synthetases
aminoacyl-tRNA synthetase
enzymes that facilitate specificity between amino acid and tRNA… there is one of these enzymes for each of the 20 amino acids… each enzyme recognizes one amino acid and their respective tRNA
amino acids are recognized by
their different shapes
tRNAs are recognized by
their differing sequences (base pairing sequences)
translation step 2. initiation
a) IF-3 … b) mRNA binds to 30S … c) IF-2 … d) IF-1
IF-3
1st step of initiation – prevents subunit from joining to one another
mRNA binds to 30S
2nd step of initiation – using Shine-Dalgarno (consensus sequence found directly upstream of true start codon) and 16S rRNA
IF-2
3rd step of initiation – allows initiator tRNA with N-fMet to bind to start codon
IF-1
4th step of initiation – allows large subunit to bind to small subunit, securing initiator tRNA in P site
translation step 3. elongation
a) EF-Tu-GTP … b) EF-Ts … c) Peptidyl transferase … d) EF-G-GTP
EF-Tu-GTP
1st step of elongation – escorts the next tRNA into A site… only permits correct tRNA to be placed into A site… (proofreading enzyme, then after fulfilling proofreading, GTP -> GDP)
EF-Ts
2nd step of elongation – recycles EF-Tu-GDP to EF-Tu-GTP (GDP -> GTP)
peptidyl transferase
3rd step of elongation – polypeptide on P site tRNA is transferred to amino acid on A site tRNA, catalyzed by large subunit rRNA
EF-G-GTP
4th step of elongation – “translocation”… pushes ribosome one codon down mRNA 5’ -> 3’… the tRNA that occupied A site is now in P site (empty tRNA had been moved to E site -> discarded), so A site is available to receive new tRNA (aminoacyl tRNA)
IF
initiation factor
EF
elongation factor
polypeptide is formed
N -> C direction
translation step 4. elongation
a) stop codon moves into A site … b) release factors (RF1/RF2, RF3) bind to stop codon and ribosome … c) release factors promote cleavage of polypeptide from tRNA in P site … d) GTP is hydrolyzed to GDP by RF3 and promotes: release of empty tRNA, release of mRNA, and dissociation of ribosomal subunits
prokaryotic ribosome
two subunits: 50S and 30S (large and small)… contains three sites that can be occupied by tRNAs (exit stage (E), peptidyl site (P), aminoacyl site (A))
exit site (E)
where empty tRNA leaves
peptidyl site (P)
where peptide chain grows from tRNA connection
aminoacyl site (A)
where tRNAs (charged tRNAs – aminoacyl tRNAs) come into ribosome
eukaryotic translation differences
ribosomal subunit sizes (40S and 60S in eukaryotes)… initiation: no Shine-Dalgarno sequence, 5’ cap recognized by small subunit and cap-binding proteins, poly-A tail binds to initiation complex via tail-binding proteins, and more initiation factors involved
control of gene expression
regulation of chromatin structure, transcription, mRNA processing, mRNA stability, translation, and posttranslational modification
prokaryotic gene regulation
genes with related function are clustered together under the control of a single promoter…. all genes are transcribed into one polycistronic mRNA
polycistronic mRNA
single mRNA molecule with multiple genes
operon
group of bacterial structural genes transcribed together… contains a promoter and operator
operon components
structural genes, regulator gene, operator
structural genes
multiple genes transcribed into one mRNA; under control of one promoter… encode proteins that are used in metabolism or biosynthesis or that play a structural role in cell
regulator gene
technically not considered part of operon as it is under control of separate promoter… transcribed and translated into regulator protein and influences transcription of structural genes… genes whose products (either RNA or proteins) interact with other DNA sequences and affect transcription/translation of those sequences
regulator protein
have domains (each with characteristic structure called a “motif”) that are responsible to binding to DNA… binds to operator
operator
bound by regulator protein… often overlaps 3’ end of promoter and 5’ end of structural genes
binding of operator by regulator protein
inhibits binding of RNA polymerase to promoter, and thus preventing transcription
regulatory elements
affect expression of sequences to which they are physically linked
positive control
process that stimulate gene expression… regulator protein is an activator that stimulates transcription
negative control
processes that inhibit gene expression… regulator protein is a repressor that inhibits transcription
lac operon
first operon to be investigated in great detail, researched by Jacob and Monod in 1961
lactose
major carbohydrate found in milk… can be metabolized by E.coli in mammalian gut
lactose metabolic steps
- transported across cell membrane by permease… 2. converted into allolactose, galactose, and glucose by beta-galactosidase
lac operon components
structural genes: lacZ (beta-galactosidase), lacY (permease), lacA (transacetylase)… regulator gene: lacI… promoter: lacP… operator: lacO
coordinate induction
simultaneous synthesis of several proteins, stimulated by the inducer molecule
negative inducible operon
regulator protein is a repressor .. transcription is by default off (inhibited) and must be turned on (induced)
lac operon (negative inducible)
regulator protein is a repressor (negative operon)–> repressor binds to operator and physically blocks binding of RNA polymerase –> transcription is by default off (inhibited) and must be turned on (induced) (inducible operon) –> an inducer (allolactose) binds to repressor and changes its shape to make it inactive… repressor is prevented from binding the operator so RNA polymerase is able to bind promoter and transcription can proceed
allosteric protein
proteins that change shape upon binding to another molecule… such as an active regulator protein that changes shape after binding with an inducer
repressible operon
transcription is normally on and must be turned off
negative operon
regulator protein is a repressor
inducible operon
transcription is normally off and must be turned on
lac operon when lactose is absent
- active repressor binds to operator… 2. RNA polymerase cannot bind to promoter… 3. transcription prevented
lac operon when lactose is present
- some lactose is converted into allolactose… 2. allolactose inactivates repressor… 3. repressor cannot bind operator… 4. RNA polymerase is able to bind to promoter… 5. structural genes are expressed… 6. lactose is metabolized
lac paradox solution
basal level of transcription allows minimal production of permease and beta-galactosidase to allow lactose conversion into allolactose
lac mutation
partial diploid: merozygote (normal haploid genome with plasmid containing an extra lac operon copy) / E.coli strain with an extra copy of lac operon while the remainder of genome is haploid
constitutive
structural genes that encode essential cellular functions and are expressed continually… such as functional lac enzymes being produced whether lactose is present or not
wild type lac operon
lacI+ lacP+ lacO+ lacZ+ lacY+ / lacI+ lacP+ lacO+ lacZ+ lacY+
[no beta-galactosidase and permease without allolactose… yes beta-galactosidase and permease with allolactose]
lacI- mutation
repressor mutation, the repressor is inactive
lacIs mutation
superrepressor mutation… prevents transcription even in presence of lactose because it has an altered inducer binding site, which means that the inducer can never bind to repressor to release the operator… dominant over I+
lacOc mutation
operator mutation… repressor cannot bind to the operator so transcription will always occur with/without lactose (constitutive)… dominant over lacO+… only affects genes on same chromosome (cis acting)
lacP- mutation
promoter mutation… RNA polymerase cannot bind promoter so transcription can never occur… is cis acting
types of operons
negative inducible… negative repressible… positive inducible… positive repressible
negative repressible operon
transcription is turned on by default and must be turned off… regulator protein is produced in an inactive form so it cannot bind operator and allows structural genes to be expressed… a corepressor binds to repressor to allosterically alter its shape to an active form… repressor can now bind to operator and halt transcription
operon is turned off when needed
product U is present at high concentration… enzymatic pathway should be halted… enzymes G, H, and I are not produced
positive inducible operon
regulator proteins are activators… transcription is off by default with the regulator protein initially produced in inactive form… with precursor present: precursor binds to regulator protein -> activates regulator protein -> regulator protein binds DNA but does not bind to operator -> transcription is activated -> structural genes are expressed
positive repressible operon
transcription is on by default with regulator protein being produced in active form… if end product is present at high levels: end product binds to activator -> regulator protein is inactivated -> regulator protein does not bind DNA -> transcription is halted and structural genes are not expressed
catabolite repression
when glucose is present then genes controlling the metabolism of other sugars are repressed…. results from positive (activator) control in response to absence of glucose (catabolite activator protein binds upstream of lac structural promoter in presence of cyclic AMP)… cAMP levels are high when glucose levels are low… works in concert with negative inducible control (cell is more responsible to changes in environment
when glucose levels are low
cAMP levels are high -> cAMP binds CAP -> cAMP-CAP recruits RNA polymerase to promoter -> lac structural genes are expressed
when glucose levels high
cAMP levels are low-> cAMP does not bind CAP -> unactivated CAP does not recruit RNA polymerase -> lac structural genes are expressed at a very low level
population
group of interbreeding members of a single species
gene pool
set of genetic information carried by the members of a sexually reproducing population
allelic frequency
frequency of an allele in a gene pool… p= f(A) = 2nAA + nAa / 2N and q = f(a) = 2naa + nAa / 2N… divide by number of alleles in gene pool
genotypic frequency
frequency of a genotype in a population… f(AA) = # AA individuals / N and f(Aa) = # Aa individuals / N and f(aa) = # aa individuals / N… divide by number of individuals
hardy-weinberg law
if population is large (genetic drift); no natural selection; random mating (selection); no mutation; and no migration………… then 1. allelic frequencies in gene pool of a population will not change from one generation to the next AND 2. genotypic frequencies will stabilize after one generation of the previous condition to p^2, 2pq, and q^2
Hardy-Weinberg equilibrium
p^2 + 2pq + q^2… p^2 = frequency of AA genotype, 2pq = frequency of Aa genotype, and q^2 = frequency of aa genotype… p = frequency of A allele and q = frequency of a allele
Hardy-Weinberg Details
“large” population is any population that is not small… random mating implies that each genotype mates in proportion to its frequency… each H-W applies to one locus (assumptions must hold true for only the locus being studied… a population may be in equilibrium for some loci but not others)
implications of Hardy-Weinberg
- population cannot “evolve” if it meets H-W assumptions– evolution requires changes in allelic frequency; random mating alone does not bring about changes in frequencies, so other factors must be present in order for a population to “evolve”… 2. when in H-W equilibrium, genotypic frequencies are determined by allelic frequencies– heterozygote frequency is greatest when allelic frequency is 0.5 (but never exceeds that frequency); when an allele is at low frequency, most alleles will be found in heterozygotes
eugenics
the science of improving a human population by controlled breeding to increase the occurrence of desirable heritable characteristics… the breeding of humans to favor certain traits
Using Hardy-Weinberg
used to estimate the frequency of alleles and genotypes in a population in genetic equilibrium
extension of H-W: multiple alleles
allelic frequencies: f(A’) = p , f(A’’) = q , f(A’’’) = r … genotypic frequencies: p^2 = f(A’A’) , 2pq = f(A’A’’), , q^2 = f(A’‘A’’) , 2pr = f(A’A’’’) , 2qr = (A’‘A’’’) , r^2 = f(A’'’A’’’)
extension of H-W: X-linked alleles
allelic frequencies: f(X’) = p , f(X’’) = q … genotypic frequencies: f(X’X’) = p^2 , f(X’X’’) = 2pq , f(X’‘X’’) = q^2 , f(X’Y) = p , f(X’‘Y) = q
chi-square test with Hardy-Weinberg Equilibrium
- Ho and Ha (____ gene with ___ alleles is/is not in Hardy-Weinberg equilibrium for this population)… 2. calculate p and q (and r if applicable)… 3. calculate expected values (using appropriate frequency products, ie p^2, 2pq, q^2, etc)… 4. calculate chi-square… 5. compare to p-table values… 6. conclusion (there is between a ____ and ____ % that any differences between observed and expected values are due to random chance and ____ and ____ % that any differences between observed and expected values are due to some actual physical factor. the p-value is smaller/larger than the significant threshold of p = 0.05, so we reject/fail to reject the Ho and reject/fail to reject the Ha)
selection
differential reproduction of genotypes based on fitness (relative reproductive success of an individual)… does not produce new alleles (but allows some alleles to be passed to descendants at a greater frequency)… does not act on individual genes (selects combinations of genes)…. types: Natural and Artificial
natural selection
not human directed, environmentally controlled
artificial selection
human directed/controlled
selection and nonrandom mating
affects the probability that certain alleles combine to form genotypes– alters genotypic frequencies
positive assortative mating
tendency for similar individuals to mate
negative assortative mating
tendency for dissimilar individuals to mate
inbreeding
preferential mating between related individuals (positive assortative mating for relatedness)… affects all genes… leads to departure from H-W equilibrium, increased proportion of homozygotes, decreased proportion of heterozygotes, and increased incidence of genetic diseases
outbreeding
negative assortative mating for unrelatedness
outcrossing
avoidance of mating between related individuals
genetic rescue
introduction of new genetic variation into an inbred population
identical by descent
two alleles of a homozygote are descended from same ancestral allele (inbreeding)
identical by state
two alleles of a homozygote are identical in structure and function but are from two different copies in ancestors (outbreeding)
inbreeding coefficient (F)
measure of probability that two alleles are identical by descent… varies from 0 to 1 (with 0 indicating random mating and 1 indicating that all alleles are identical by descent)… if F=1 then self-fertilization or cloning has occurred
effect of inbreeding on H-W
increase in homozygotic frequencies– f(AA) = p^2 + Fpq ; f(aa) = q^2 + Fpq … decrease in heterozygote frequencies– f(Aa) = 2pq - 2Fpq
inbreeding depression
increased appearance of lethal and deleterious phenotypes as a result of inbreeding… increased numbers of homozygotes will lead to a greater proportion of affected individuals… inbreeding is harmful to fitness of populations
migration (aka gene flow)
movement of some individuals from one population to another, resulting in a change in allelic frequency
calculating allelic frequencies after migration
q’II = qIm + qII(1-m) [where q’II = little allele frequency in mixed population ; qI = little allele frequency in population 1 (migrant population) ; m = fraction of mixed population that came from the migrant population ; qII = little allele frequency in population 2] OR p’II = pIm + pII(1-m) [where p’II = big allele frequency in mixed population ; pI = big allele frequency in population 1 (migrant population) ; m = fraction of mixed population that came from the migrant population ; pII = big allele frequency in population 2] …. Δq = m(qI-qII) is the change in little allele frequency OR Δp = m(pI-pII) is the change in big allele frequency
effects of migration
prevents genetic divergence between populations (reduces differences in allele frequencies between populations and tends to make populations more alike)… increased genetic variation within populations (spreads new alleles between different populations)
mutation
sudden heritable change in genetic material, resulting in some new alleles/genetic variants… usually detrimental and recessive in nature… types: nonrecurrent (tends to only occur once) or recurrent (can occur in future generations, nonreversible or reversible)… does not have a large effect on allele frequencies… mutated allele is maintained in gene pool once it is enriched in population due to selection… slow accumulation of mutations is what provides genetic variability
genetic drift
when population size is limited– gametes that randomly unite to form individuals of next generation carry only a sample of alleles present in parental gene pool, so proportions in sampled gametes can differ from those of parental generation due to random chance… sampling effects can lead to deviations from expected frequencies (sampling error)… can predict only the magnitude of change because it is a dispersive process
genetic drift effects
changes allelic frequencies within a population… reduces genetic variation within a population (may lead to fixation of an allele)… increases genetic diversity between populations
effective population size (Ne)
population of individuals that contribute genes to next generation… number of breeding adults
causes of genetic drift
- founder effects– establishment of an isolated population by a small number of individuals (genes carried by descendants are derived from few alleles from founders)… 2. genetic bottleneck– develops when a population undergoes a drastic reduction in size (descendants will be genetically similar because they have alleles that were carried by a few survivors of bottleneck)
