CRYSTALLOGRAPHY OF BRAIN NITRIC OXIDE SYNTHASE REDUCTASE

点击翻译按钮获取中文摘要

We aim to determine the atomic structure of the 70kD reductase domain of nitric oxide synthase from the brain. Nitric oxide (NO) is an important molecule in intercellular signalling. Nitric oxide synthase (NOS) is a large polypeptide, responsible for the production of NO. The crystal structure of one half of inducible nitric oxide synthase (the 70kD oxygenase domain, using data collected at SSRL) has been determined, but the structure of the 70kD reductase domain remains unknown. The reductase domain of nitric oxide synthase binds three redox active cofactors: FMN, FAD, and NADPH. These constitute the first three electron carriers in the electron transport chain that results in production of NO. At present the structural mechanism by which these cofactors supply electrons to the heme group of the oxygenase domain is unknown. We have obtained two crystal forms of brain NOS reductase domain (bNOSr). On our laboratory X-ray source, these crystals seldom diffract to better than 6 E resolution; however, preliminary tests at synchrotron X-ray sources have shown diffraction to 2.7 E. Therefore, synchrotron radiation is necessary to obtain X-ray diffraction data to reasonable resolution for structure determination. We are confident that with improved crystallization conditions, this limit can be further improved. We have obtained preliminary phasing information using a homologous molecular replacement search model, but we require higher quality diffraction data in order to complete the project. Our goal is to collect high quality diffraction data for bNOSr in the native form and in complex with inhibitors.