Crystal Structure of the RIM2 C2A-Domain at 1.4 Å Resolution

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• 作者：Han Dai ; Diana R. Tomchick ; Jesú ; s Garc&iacute ; a ; Thomas C. S&uuml ; dhof ; Mischa Machius ; and ; and Josep Rizo
• 刊名：Biochemistry
• 出版年：2005
• 出版时间：October 18, 2005
• 年：2005
• 卷：44
• 期：41
• 页码：13533 - 13542
• 全文大小：1188K
• 年卷期：v.44,no.41(October 18, 2005)
• ISSN：1520-4995

文摘

RIMs are large proteins that contain two C₂-domains and are localized at presynaptic activezones, where neurotransmitters are released. RIMs play key roles in synaptic vesicle priming and regulationof presynaptic plasticity. A mutation in the RIM1 C₂A-domain has been implicated in autosomal dominantcone-rod dystrophy (CORD7). The RIM C₂A-domain does not contain the full complement of aspartateresidues that commonly mediate Ca²⁺ binding at the top loops of C₂-domains, and has been reported tointeract with SNAP-25 and synaptotagmin 1, two proteins from the Ca²⁺-dependent membrane fusionmachinery. Here we have used NMR spectroscopy and X-ray crystallography to analyze the structure andbiochemical properties of the RIM2 C₂A-domain, which is closely related to the RIM1 C₂A-domain. Wefind that the RIM2 C₂A-domain does not bind Ca²⁺. Moreover, little binding of the RIM2 C₂A-domainto SNAP-25 and to the C₂-domains of synaptotagmin 1 was detected by NMR experiments, suggestingthat as yet unidentified interactions of the RIM C₂A-domain mediate its function. The crystal structure ofthe RIM2 C₂A-domain using data to 1.4 Å resolution reveals a mages/gifchars/beta2.gif" BORDER=0 ALIGN="middle">-sandwich that resembles those observedfor other C₂-domains, but exhibits a unique dipolar distribution of electrostatic charges whereby one edgeof the mages/gifchars/beta2.gif" BORDER=0 ALIGN="middle">-sandwich is highly positive and the other edge is highly negative. The location of the mutationsite implicated in CORD7 at the bottom of the domain and the pattern of sequence conservation suggestthat, in contrast to most C₂-domains, the RIM C₂A-domains may function through Ca²⁺-independentinteractions involving their bottom face.