Independently Melting Modules and Highly Structured Intermodular Junctions within Complement Receptor Type 1

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• 作者：Marina D. Kirkitadze ; Malgorzata Krych ; Dusan Uhrin ; David T. F. Dryden ; Brian O. Smith ; Alan Cooper ; Xuefeng Wang ; Richard Hauhart ; John P. Atkinson ; and Paul N. Barlow
• 刊名：Biochemistry
• 出版年：1999
• 出版时间：June 1, 1999
• 年：1999
• 卷：38
• 期：22
• 页码：7019 - 7031
• 全文大小：221K
• 年卷期：v.38,no.22(June 1, 1999)
• ISSN：1520-4995

文摘

A segment of complement receptor type 1 (CR1) corresponding to modules 15-17 wasoverexpressed as a functionally active recombinant protein with N-glycosylation sites ablated by mutagenesis(referred to as CR1~15-17^-). A protein consisting of modules 15 and 16 and another corresponding tomodule 16 were also overexpressed. Comparison of heteronuclear nuclear magnetic resonance (NMR)spectra for the single, double, and triple module fragments indicated that module 16 makes more extensivecontacts with module 15 than with module 17. A combination of NMR, differential scanning calorimetry,circular dichroism, and tryptophan-derived fluorescence indicated a complex unfolding pathway forCR1~15-17^-. As temperature or denaturant concentration was increased, the 16-17 junction appearedto melt first, followed by the 15-16 junction, and module 17 itself; finally, modules 15 and 16 becamedenatured. Modules 15 and 16 adopted an intermediate state prior to total denaturation. These results arecompared with a previously published study [Clark, N. S., Dodd, I, Mossakowska, D. E., Smith, R. A.G., and Gore, M. G. (1996) Protein Eng. 9, 877-884] on a fragment consisting of the N-terminal threeCR1 modules which appeared to melt as a single unit.