Mutations & Gene Regulation

Question 1

(a) mRNA from coding strand = same sequence with U for T: AUG-GAG-GUU-CAU-UUC-GGA-UAA → Met–Glu–Val–His–Phe–Gly–Stop (peptide: Met-Glu-Val-His-Phe-Gly). (1 mark reading frame, 1 correct AAs, 1 stop identified)

(b) (3 marks each = 9)

• Mutation A (pos6 G→A): codon2 GAG→GAA. (i) substitution; (ii) GAA=Glu = same as GAG=Glu → silent (synonymous); (iii) peptide unchanged: Met-Glu-Val-His-Phe-Gly. Marks: type 1, consequence 1, peptide 1.
• Mutation B (pos8 T→A): codon3 GTT→GAT... check — positions: codon3 = pos7,8,9 = G,T,T. Pos8 T→A gives GAT. GAT not in given table; use standard code GAT=Asp. (i) substitution; (ii) Val→Asp = missense; (iii) Met-Glu-Asp-His-Phe-Gly. Accept missense reasoning; award full if student notes amino acid change. Marks: type 1, consequence 1, peptide 1.
• Mutation C (A inserted after pos3): sequence becomes ATG-AGA-GGT-TCA-TTT-CGG-ATA-A. (i) insertion; (ii) frameshift — all downstream codons altered; (iii) reading: AGA=Arg, GGT=Gly, TCA=Ser, TTT=Phe, CGG=Arg, ATA=Ile... no early stop in this fragment → Met-Arg-Gly-Ser-Phe-Arg-Ile (runs on). Marks: type 1, frameshift 1, altered downstream reading 1. (Full credit for correctly showing frame shift disrupts all downstream codons.)

(c) (6 marks)

def classify_substitution(wild_codon, mut_codon):
    w = CODON_TABLE[wild_codon]
    m = CODON_TABLE[mut_codon]
    if w == m:            # test 1
        return "silent"
    if m == "Stop":       # test 2
        return "nonsense"
    return "missense"     # test 3 (default)

Ordered tests: (1) same amino acid → silent; (2) mutant codon is stop → nonsense; (3) otherwise different amino acid → missense. Why order matters: the silent check must come first (identical residues); the nonsense check must precede missense because a stop codon is technically a "different product" and would otherwise be mislabelled missense. (function 3, correct order stated 2, justification 1.)

(d) (4 marks) Sickle-cell: $\beta$-globin codon 6 GAG→GTG, changing Glu→Val. It is missense because a single base substitution replaces one amino acid with a different amino acid (not the same → not silent) and does not create a stop codon (→ not nonsense). The Val substitution creates a hydrophobic patch causing HbS polymerisation. (codon change 1, Glu→Val 1, missense justification 2.)

Question 2

(a) (4) Inducible: normally OFF (repressor bound), turned ON by an inducer — default state OFF (lac). Repressible: normally ON, turned OFF when corepressor binds — default state ON (trp). (2 marks each system.)

(b) (6) Transcription needs repressor OFF (lactose present, $L=1$) AND CAP active (glucose absent, $G=0$): $T_{lac} = L \land \lnot G$ Truth table:

(expression 2, table 4 rows correct 4 — deduct per wrong row.) Only high expression when lactose present and glucose absent.

(c) (4) $T_{trp} = \lnot W$.

Corepressor = tryptophan itself; it binds the inactive repressor, changing its conformation so it can bind the operator and block transcription (feedback shutoff of trp synthesis). (expression 1, table 1, corepressor role 2.)

(d) (6) Constitutive expression mutations:

• Operator mutation ($O^c$): repressor cannot bind operator → transcription always on. cis-acting — a mutated DNA site affects only the adjacent operon on the same molecule. (3)
• Repressor gene mutation ($lacI^-$): non-functional repressor protein → cannot repress. trans-acting — the repressor is a diffusible protein product acting on any operon copy. (3) (Justify cis = DNA site/same molecule; trans = diffusible product.)

Question 3

(a) (5) Germline mutations occur in gametes/germ cells → heritable, present in every cell of offspring. Somatic mutations occur in body cells → not passed to offspring, affect only the mutant clone (e.g., cancers). DNA methylation marks in the germline are largely erased and re-established during gametogenesis, so most somatic epigenetic silencing is not inherited; however, imprinted/incompletely reset methylation can be transgenerationally inherited, unlike a somatic methylation change confined to one tissue. (germline vs somatic 3, epigenetic reprogramming/heritability 2.)

(b) (5) miRNA/RNAi: miRNA (or siRNA) is processed by Dicer, loaded into RISC (Argonaute); the guide strand base-pairs with complementary sequence in the target mRNA 3'UTR → translational repression or mRNA cleavage/degradation. This acts post-transcriptionally (regulates existing mRNA). DNA methylation acts transcriptionally (prevents mRNA from being made by recruiting repressive chromatin). (mechanism 3, level contrast 2.)

(c) (i) $N(t) = N_0 e^{-kt}$. Fraction remaining $= e^{-0.35\times4} = e^{-1.4} \approx 0.2466$ (≈24.7%); $N(4)\approx 1973$ molecules. (equation 2, value 2.) (ii) $t_{1/2} = \dfrac{\ln 2}{k} = \dfrac{0.6931}{0.35} \approx 1.98\ \text{h}$. (2.) (iii) $\dfrac{dN}{dt} = s - kN$. Steady state $dN/dt=0 \Rightarrow N_{ss} = s/k = 1400/0.35 = 4000$ molecules. (ODE 1, N_ss 1.)

[
 {"claim":"Fraction remaining at t=4h is exp(-1.4)≈0.2466","code":"import sympy as sp\nk=sp.Rational(35,100); t=4\nfrac=sp.exp(-k*t)\nresult = abs(float(frac)-0.2465969)<1e-4"},
 {"claim":"N(4)≈1973 molecules for N0=8000","code":"import sympy as sp\nN=8000*sp.exp(-sp.Rational(35,100)*4)\nresult = abs(float(N)-1972.78)<0.5"},
 {"claim":"Half-life = ln2/0.35 ≈ 1.98 h","code":"import sympy as sp\nth=sp.log(2)/sp.Rational(35,100)\nresult = abs(float(th)-1.98046)<1e-3"},
 {"claim":"Steady state N_ss = s/k = 4000","code":"import sympy as sp\nN=sp.symbols('N'); s=1400; k=sp.Rational(35,100)\nsol=sp.solve(s-k*N,N)[0]\nresult = sol==4000"},
 {"claim":"lac transcription truth: only L=1,G=0 gives 1","code":"import sympy as sp\ndef T(L,G): return int(L==1 and G==0)\nrows=[T(0,0),T(0,1),T(1,0),T(1,1)]\nresult = rows==[0,0,1,0]"}
]