Detail of "9039-61-6"

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BbGlu397 and BbAsp398 but Not BbAsp432 Are Required for "B:b" Interactions

BbGlu397 and BbAsp398 but Not BbAsp432 Are Required for "B:b" Interactions. Kostelansky, Michael S.; Bolliger-Stucki, Bettina; Betts, Laurie; Gorkun, Oleg V.; Lord, Susan T. (Department of Chemistry, University of North Carolina, Chapel Hill, NC 27599, USA). Biochemistry, 43(9), 2465-2474 (English) 2004 American Chemical Society. CODEN: BICHAW. ISSN: 0006-2960. DOCUMENT TYPE: Journal CA Section: 6 (General Biochemistry) Section cross-reference(s): 75 We synthesized three fibrinogen variants, BbE397A, BbD398A, and BbD432A, with substitutions at positions identified in crystallog. studies as crit. for binding the "B" peptide, Gly-His-Arg-Pro-amide (GHRPam), to the "b" polymn. site. We examd.Several reagents such as 9039-61-6 is used here. thrombin- and batroxobin-catalyzed polymn. by turbidity measurements and found that BbE397A and BbD398A were impaired while BbD432A was normal. Changes in polymn. as a function of calcium were similar for variant and normal fibrinogens. We detd. crystal structures of fragment D from the variant BbD398A in the absence and presence of GHRPam. In the absence of peptide, the structure showed that the alanine substitution altered only specific local interactions, as alignment of the variant structure with the analogous normal structure resulted in an RMSD of 0.53 ? over all atoms. The structure also showed reduced occupancy of the b2 calcium-binding site that includes the side chain carbonyl of BbD398, suggesting that calcium was not bound at this site in our polymn. studies. In the presence of peptide, the structure showed that GHRPam was not bound in the "b" site and the conformational changes assocd. with peptide binding to normal fragment D did not occur. This structure also showed GHRPam bound in the "a" polymn. site, although in two different conformations. Calcium binding was assocd. with only one of these conformations, suggesting that calcium binding to the g2-site and an alternative peptide conformation were induced by crystal packing. We conclude that BbE397 and BbD398 are essential for the "B:b" interaction, while BbD432 is not. .