期刊
JOURNAL OF BIOLOGICAL CHEMISTRY
卷 287, 期 39, 页码 32897-32912出版社
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
DOI: 10.1074/jbc.M112.354365
关键词
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资金
- National Institutes of Health [CA099835, CA118595, AI076961, AI072106, AI068730, AI081982, AI2008031, GM037684, GM020501, GM066170, GM093325, NS070899, RR029388]
- University of California IUCRP Program, Biogen Idec [UC10591]
We have applied hydrogen-deuterium exchange mass spectrometry, in conjunction with differential scanning calorimetry and protein stability analysis, to examine solution dynamics of the integrin alpha 1 I domain induced by the binding of divalent cations, full-length type IV collagen, or a function-blocking monoclonal antibody. These studies revealed features of integrin activation and alpha 1I-ligand complexes that were not detected by static crystallographic data. Mg2+ and Mn2+ stabilized alpha 1I but differed in their effects on exchange rates in the alpha C helix. Ca2+ impacted alpha 1I conformational dynamics without altering its gross thermal stability. Interaction with collagen affected the exchange rates in just one of three metal ion-dependent adhesion site (MIDAS) loops, suggesting that MIDAS loop 2 plays a primary role in mediating ligand binding. Collagen also induced changes consistent with increased unfolding in both the alpha C and allosteric C-terminal helices of alpha 1I. The antibody AQC2, which binds to alpha 1I in a ligand-mimetic manner, also reduced exchange in MIDAS loop 2 and increased exchange in alpha C, but it did not impact the C-terminal region. This is the first study to directly demonstrate the conformational changes induced upon binding of an integrin I domain to a full-length collagen ligand, and it demonstrates the utility of the deuterium exchange mass spectrometry method to study the solution dynamics of integrin/ligand and integrin/metal ion interactions. Based on the ligand and metal ion binding data, we propose a model for collagen-binding integrin activation that explains the differing abilities of Mg2+, Mn2+, and Ca2+ to activate I domain-containing integrins.
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