EVIDENCE FOR A POST-TRANSLATIONAL MODIFICATION, ASPARTYL ALDEHYDE

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 在氧合光合作用中，光系统II(PSII)进行水的氧化和叶绿体对苯二酚的还原。三个PSII亚基含有共价结合胺和苯肼的反应性基团。有人提出，这些反应性基团是含有羰基的、共或翻译后修饰的氨基酸(Ouellette等人。程序娜塔莉。阿卡德。SCI。U.S.A.1998、95、2204和Anderson等人。J.Biol.化学。2000、275、4920)。为了鉴定其中一个PSII亚基(CP47)的修饰氨基酸残基，进行了串联质谱分析。修饰的残基被生物素-LC-酰肼或生物细胞素肼亲和标记，这两种标记是已知的标记羰基。用变性凝胶电泳法分离亲和标记的亚基，并对其进行亲和纯化和串联质谱分析。本程序鉴定了一种肼标记的多肽，其序列为XKEGR。这一结果得到了肽图和甲胺标记的定量结果的支持。基因序列和这些串联数据预测，用酰肼试剂标记的第一个氨基酸X是天冬氨酸的修饰形式。在这些数据的基础上，我们认为CP47亚基的D348在翻译后或共翻译后被修饰，得到了一个新的氨基酸侧链，天冬氨酸醛。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. In oxygenic photosynthesis, photosystem II (PSII) carries out the oxidation of water and reduction of plastoquinone. Three PSII subunits contain reactive groups that covalently bind amines and phenylhydrazine. It has been proposed that these reactive groups are carbonyl-containing, co- or post-translationally modified amino acids (Ouellette et al. Proc. Natl. Acad. Sci. U.S.A. 1998, 95, 2204 and Anderson et al. J. Biol. Chem. 2000, 275, 4920). To identify modified amino acid residues in one of the PSII subunits (CP47), tandem mass spectrometry was performed. Modified residues were affinity-tagged with either biotin-LC-hydrazide or biocytin hydrazide, which are known to label carbonyl groups. The affinity-tagged subunit was isolated by denaturing gel electrophoresis, and tryptic peptides were then subjected to affinity purification and tandem mass spectrometry. This procedure identified a hydrazide-labeled peptide, which has the sequence XKEGR. This result is supported by quantitative results acquired from peptide mapping and methylamine labeling. The gene sequence and these tandem data predict that the first amino acid, X, which is labeled with the hydrazide reagent, is a modified form of aspartic acid. On the basis of these data, we propose that D348 of the CP47 subunit is post- or co-translationally modified to give a novel amino acid side chain, aspartyl aldehyde.