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When performing site‑directed mutagenesis to test predictions about which residues are essential for protein function, which amino acid substitution would disrupt protein structure the most.

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tushargupta0691

Premature stop codon mutations are the most harmful ones because they are more likely to cause a complete loss of function the earlier in the sequence they occur.

The most likely structural changes occur when (conserved) Gly is changed to a non-Gly residue and when Pro is changed because these changes narrow the permissible range of main-chain torsion angles. Cys mutations are also likely to have an impact on extracellular proteins: losing a disulfide bond may make your protein less stable, and unpaired Cys residues may cause crosslinking if the side chain is exposed.

The impact of conservative substitutes will probably be less severe than that of non-conservative ones. A multi-sequence alignment can also help you determine which property—such as size, charge, hydrophobicity, or hydrogen bonding—is crucial in this situation.

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