ADP-RIBOSYLATION CYCLES

单腺苷二磷酸核糖基化是蛋白质翻译后的修饰 它参与细菌毒素的作用， 细胞过程。 精氨酸、半胱氨酸和天门冬氨酸特异性ADP- 核糖基转移酶已经在动物组织中被鉴定。 NAD：精氨酸 ADP-核糖基转移酶和ADP-核糖基精氨酸水解酶，其催化 假定的ADP-核糖基化循环的正向和反向臂具有 在哺乳动物组织中被发现。 ADP-核糖基精氨酸水解酶， 从火鸡红细胞和大鼠脑组织中分离纯化了其cDNA， 从后者的组织。 在精氨酸特异性ADP的情况下， 从兔骨骼肌中部分纯化核糖基转移酶 膜报道。 为了确定这种酶的结构， 纯化、微测序和克隆。 兔骨骼肌精氨酸特异性ADP核糖基转移酶， 通过连续五步色谱法纯化约215，000倍。 对 HPLC纯化的胰蛋白酶肽的氨基酸序列的基础， 合成简并寡核苷酸引物，并用于 通过基于聚合酶链式反应（PCR）的程序来产生cDNA。 一 基于PCR产生的序列，使用特异性探针来筛选 兔骨骼肌文库。 一个复合序列，从 文库筛选和cDNA 5 ′端的快速扩增， 包含一个981碱基对的开放阅读框架，编码一个36，134-Da的 蛋白 用表达载体转化的大肠杆菌细胞 含有转移酶特异性序列表达的转移酶活性。 转移酶特异性寡核苷酸探针识别4-乙酰转移酶mRNA， 主要在兔骨骼肌和心肌中表达。 没有 肌肉推导氨基酸序列的扩展相似性 转移酶和几种ADP-核糖基化细菌毒素。

Mono-ADP-ribosylation is a posttranslational modification of proteins which is involved in the action of bacterial toxins and in the regulation of cellular processes. Arginine-, cysteine- and aparagine-specific ADP- ribosyltransferases have been identified in animal tissues. NAD:arginine ADP-ribosyltransferase and ADP-ribosylarginine hydrolases, which catalyze the forward and reverse arms of a putative ADP-ribosylation cycle, have been identified in mammalian tissues. ADP-ribosylarginine hydrolase was purified from turkey erythrocytes and rat brain and its CDNA was cloned from the latter tissue. In the case of the arginine-specific ADP- ribosyltransferase, partial purification from rabbit skeletal muscle membranes was reported. To define the structure of this enzyme, it was purified, microsequenced, and cloned. Rabbit skeletal muscle arginine-specific ADP-ribosyltransferase was purified about 215,000-fold by sequential five-step chromatography. On the basis of the amino acid sequences of HPLC-purified tryptic peptides, degenerate oligonucleotide primers were synthesized and used in a polymerase chain reaction (PCR)-based procedure to generate CDNA. A specific probe, based on PCR-generated sequence, was used to screen a rabbit skeletal muscle library. A composite sequence, obtained from library screening and rapid amplification of the 5' end of the CDNA, contained a 981-base-pair open reading frame, encoding a 36,134-Da protein. Escherichia coli cells transformed with an expression vector containing transferase-specific sequence expressed transferase activity. A transferase-specific oligonucleotide probe recognized a 4-kilobase MRNA expressed primarily in rabbit skeletal and cardiac muscle. There was no extended similarity in deduced amino acid sequences of the muscle transferase and several ADP-ribosylating bacterial toxins.