【作者】 冯莲；

【导师】 李有志；

【作者基本信息】 广西大学 ， 生物化学与分子生物学， 2007， 硕士

【摘要】 以早期筛选到的一株青霉菌Penicillium janthinellum菌株GXCR为试验材料，对铜离子诱导和未诱导两种情况下所产生的胞内铜结合蛋白进行分离纯化和鉴定，为克隆与铜抗性相关的基因奠定基础。在未诱导的情况下，提取胞内蛋白，经铜亲和层析柱分离出一系列与Cu²⁺相结合的蛋白质，对其中的12个蛋白进行MALDI-TOF-MS质谱分析，分析结果提交在SWISS-PROT和NCBI数据库，发现其中的12个蛋白质与数据库的部分蛋白质相匹配，分别为谷氨酸脱氢酶、泛素连接酶E3、泛素、组氨酰-tRNA合成酶、谷氨酸脱氢酶、天冬氨酸转氨酶、Metaxin1 homolog、SHE4，其余4个为假定蛋白。目前尚未有这些蛋白质与重金属抗性相关的报道，它们在Penicillium janthinillum GXCR中抗重金属的作用还有待于进一步证实。铜离子诱导产生的铜结合蛋白经葡聚糖凝胶（Sephadex G-75）层析和离子交换（纤维素DEAE-52）层析，分离纯化出约为20kDa的铜结合蛋白纯品。经MALDI-TOF-MS分析得到的肽质量指纹谱在SWISS-PROT和NCBI数据库中搜索不到相匹配的蛋白，还需要进一步鉴定。更多还原

【Abstract】 By using Penicillium janthinellum GXCR which has been screenedbefore as material, the experiment was carried out to obtain proteins aboutcopper resistance by separated and identified copper binding proteins fromPenicillium janthinellum GXCR with copper ion inducing or without copperion inducing respectively.The proteins were extracted from Penicillium janthinellum GXCRwithout copper ions inducing. A series of copper binding proteins werepurified by copper affinity chromatography. 17 copper binding proteins wererandom selected and analysed by matrix-assisted laser desorption ionizationtime-of-flight mass spectrometry. A summary of the identified proteins arepresented. The profile of peptide mass fingerprinting of 12 proteins weresubmitted in SWISS-PROT database and NCBI database, and the resultsrevealed that they showed higher homology with thses following proteins:NADP-specific glutamate dehydrogenase, E3 ubiquitin protein ligase TOM1,ubiquitin peptide,histidyl-tRNA synthetase,NADP linked glutamate dehydrogenase,aspartate aminotransferase,metaxin 1 homolog, SHE 4 proteinand 4 hypothetical proteins. Up to now the functions of metaxin 1 homolog,SHE 4 protein, and 4 hypothetical protein are still unknow. There are noreports about these proteins response to heavy metal resistance of microbes.The function of these proteins for the heavy metal resistance need to be studying more.The copper binding protein from Penieillium janthinellum GXCR undercopper ions inducing was isolated by Sephadex G-75 and Cellulose DEAE-52.The analysis of SDS-PAGE electrophoresis revealed that the MW of purifiedcopper binding protein was approximate 20kDa.This protein was identified using peptide mass fingerprinting andmatrix-assisted laser desorption ionization time-of-flight mass spectrometry.No matched protein was found in the SWISS-PROT database and NCBIdatabase, so it is worth further studying.更多还原