Point mutations and sequence variability in proteins: Redistributions of preexisting populations -- Data Supplement - Supplemental Data -- Proceedings of the National Academy of Sciences

Supplementary material for Sinha and Nussinov (2001) Proc. Natl. Acad. Sci. USA 98 (6), 3139–3144. (10.1073/pnas.051399098)

Results
Transthyretin, HIV Proteinase, Barnase, Lactoferrin, Dehydrofolate Reductase, a -1 Antitrypsin, BP-RNase A, Adenylate Kinase, and Calmodulin.
Appendix A and Table 1 list the results, and Figs. 1 and 3 display these results when there are over four mutant structures, i.e., transthyretin, HIV proteinase, barnase, and lactoferrin. Most of the mutations are of similar types. However, as Appendix A and Table 1 show, some are of different types (three in transthyretin, three in HIV proteinase, three in barnase, one in lactoferrin, one in dihydrofolate reductase, five in a -antitrypsin, four in BP-RNase A, and two in adenylate kinase). In all cases, except for dihydrofolate reductase, the regions showing the largest deviations are in coils. In dihydrofolate reductase, the largest deviation is in a b -strand, at Ser-135, consistent with previous observations (1). However, the second-largest deviation is in a coil. The regions that deviate the most are largely, although not necessarily, around the active or binding site (Fig. 3 and Table 2). In the case of transthyretin the loops that manifest the largest deviations (Fig. 3c) fall in the region where it binds to retinol-binding protein (2). In eight out of nine cases, the highest or the second-highest deviation is at position 100–102. This position falls in the coil that connects b -strands F and G. This b -hairpin at the C terminus may be critical for amyloid formation, where the coil allows the hairpin to flip and domain swap, to form the highly stable protofibril (3). In HIV proteinase, the highest and second-highest distances are at Gly-51 or Ser-37 (Appendix A). The bound and unbound conformations of the HIV proteinase differ (4), with the flaps that form from the two b -strands forming the ceiling of the binding site. These flaps have been shown to move from an open to a closed conformation (5). Gly-51 lies in the coil of the flap region, between strands C and D, whereas Ser-37 is in the coil between strands B and G (Fig. 3d). Both positions are important for function. In barnase, in all mutants, the higher deviations are at positions 59 to 68, around Gly-40 or -104 (Appendix A, Table 1). Positions 59–68 fall between strands C and D. Position 40 is in the coil between helix B and strand C, whereas position 104 falls in the coil between strand F and the C terminus (Fig. 3e). The coils at the C terminus and between helix B and strand C form part of the binding site. The most common deviation in barnase is in the green coil, between helices A and B.

In lactoferrin in all mutants the highest deviations are at Arg-313. Lactoferrin is a 691-residue protein. However, the mutant structures 1hse, 1dsn, 1vfd, and 1vfe were solved only for the N-terminal 320 residues. The protein consists of N- and C-terminal domains. The N-terminal domain contains two lobes that have been shown to move relative to each other around the binding site (6). Pro-284 is second highest in two of the cases.

The deviations were computed for two yeast adenylate kinases (1dvr, 3aky) with respect to the one from Escherichia coli. The highest and second-highest deviations fall in the same region for both yeast adenylate kinases. For calmodulin, chicken and bovine (1ahr, 1deg) were compared with the Drosophila calmodulin (4cln). Here, calmodulin from different sources shows the effect of mutations in the same region.

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