Impaired Protofibril Formation in Fibrinogen γN308K Is Due to Altered D:D and “A:a” Interactions,

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• 作者：Sheryl R. Bowley ; Nobuo Okumura ; Susan T. Lord
• 刊名：Biochemistry
• 出版年：2009
• 出版时间：September 15, 2009
• 年：2009
• 卷：48
• 期：36
• 页码：8656-8663
• 全文大小：1096K
• 年卷期：v.48,no.36(September 15, 2009)
• ISSN：1520-4995

文摘

“A:a” knob−hole interactions and D:D interfacial interactions are important for fibrin polymerization. Previous studies with recombinant γN308K fibrinogen, a substitution at the D:D interface, showed impaired polymerization. We examined the molecular basis for this loss of function by solving the crystal structure of γN308K fragment D. In contrast to previous fragment D crystals, the γN308K crystals belonged to a tetragonal space group with an unusually long unit cell (a = b = 95 Å, c = 448.3 Å). Alignment of the normal and γN308K structures showed the global structure of the variant was not changed and the knob “A” peptide GPRP was bound as usual to hole “a”. The substitution introduced an elongated positively charged patch in the D:D region. The structure showed novel, symmetric D:D crystal contacts between γN308K molecules, indicating the normal asymmetric D:D interface in fibrin would be unstable in this variant. We examined GPRP binding to γN308K in solution by plasmin protection assay. The results showed weaker peptide binding, suggesting that “A:a” interactions were altered. We examined fibrin network structures by scanning electron microscopy and found the variant fibers were thicker and more heterogeneous than normal fibers. Considered together, our structural and biochemical studies indicate both “A:a” and D:D interactions are weaker. We conclude that stable protofibrils cannot assemble from γN308K monomers, leading to impaired polymerization.