Structural effects induced by removal of a disulfide-bridge: the X-ray structure of the C30A/C51A mutant of basic pancreatic trypsin inhibitor at 1.6 A

The X-ray structure of a variant of basic pancreatic trypsininhibitor (BPTI) has been analyzed to determine the structuralaccommodation resulting from removal of a disulfide crosslinkin a protein. The disulfide removed, Cys30–Cys51, hasbeen implicated in both the folding pathway of the protein andits overall thermal stability. In the variant studied, C30A/C51A,the disulfide cysteines were replaced by less bulky alanines.The atomic displacements observed for C30A/C51A indicate a setof concerted shifts of two segments of chain, which togethersignificantly diminish a packing defect at the site of the removedcysteine sulfur atoms. The observed structural changes are distributedasymmetrically around the sites of mutation, indicating thatthe adjacent ß-sheet is more resistant to the perturbationthan the -helix on the opposite side of the disulfide bond.The thermal parameters of groups involved in the structuralaccommodation are not significantly altered. A comparison ofthe X-ray structures reported for native BPTI determined inthree different crystal forms indicates that the magnitude ofits conformational variability exceeds that of the structuralchanges caused by the disulfide removal. This emphasizes thenecessity of using isomorphous crystal systems to determinethe relatively small effects due to mutation.