Quirin-Stricker C, Schmitt M, Egly J M, Kempf J
Eur J Biochem. 1976 Feb 2;62(1):199-209. doi: 10.1111/j.1432-1033.1976.tb10114.x.
One ribosomal protein kinase activity and 3 soluble protein kinase activities have been identified in plasma cell tumors by DEAE-cellulose chromatography. We have shown phosphorylation in vivo and in vitro of a protein fraction from the ribosomal KCl wash which we have termed 'PPx fraction'. Phosphorylation of this protein fraction has been obtained in vitro with the ribosome-associated protein kinase. We have determined for the ribosomal protein kinase the following characteristics. 1. It is an Mg2+-dependent enzyme that transfers the gamma-phosphate from ATP into phosphoseryl and phosphothreonyl residues of the substrate. 2. It has a wide substrate specificity. Like the soluble protein kinases it catalyses the phosphorylation of several proteins like histone, phosvitin, casein and ribosomal proteins but it differs from the main soluble kinases (I, II) by the fact that it catalyses specifically the phosphorylation at least of one of the ribosomal KCl wash proteins. On dodecylsulfate-polyacrylamide gels this protein has a molecular weight of approximately 90000 and it is released from ribosomes under conditions commonly employed for extraction of initiation factors. 3. The ribosome-associated protein kinase is not stimulated by the addition of cyclic adenosine 3':5'-monophosphate. 4. KCl has no effect, NaCl has a weak effect on the phosphorylation, Mn2+ and Ca2+ are inhibitors. 5. ADP has been found to be a competitive inhibitor. 6. The maximum velocity of the ribosomal protein-kinase-catalysed reaction is 0.65 nmol of 32P incorporated in the KCl wash protein per min and per mg protein. 7. The apparent Km for the ribosomal KCl wash protein as substrate is 0.71 mg/ml and the Km for ATP is 94 muM. 8. The molecular weight of the ribosomal protein kinase, estimated by electrophoresis in polyacrylamide-dodecylsulfate gels, is 60000 and corresponds probably to a catalytic subunit.
通过二乙氨基乙基纤维素色谱法，在浆细胞瘤中鉴定出一种核糖体蛋白激酶活性和三种可溶性蛋白激酶活性。我们已经证明了核糖体氯化钾洗脱液中的一种蛋白组分（我们称之为“PPx组分”）在体内和体外都能发生磷酸化。该蛋白组分在体外可被核糖体相关蛋白激酶磷酸化。我们已经确定了核糖体蛋白激酶的以下特性。1. 它是一种依赖镁离子的酶，能将ATP的γ-磷酸基团转移到底物的磷酸丝氨酸和磷酸苏氨酸残基上。2. 它具有广泛的底物特异性。与可溶性蛋白激酶一样，它能催化几种蛋白质的磷酸化，如组蛋白、卵黄高磷蛋白、酪蛋白和核糖体蛋白，但它与主要的可溶性激酶（I、II）不同，它能特异性催化至少一种核糖体氯化钾洗脱蛋白的磷酸化。在十二烷基硫酸钠-聚丙烯酰胺凝胶上，这种蛋白的分子量约为90000，并且在常用于提取起始因子的条件下从核糖体中释放出来。3. 添加环腺苷酸3':5'-单磷酸不会刺激核糖体相关蛋白激酶。4. 氯化钾无作用，氯化钠对磷酸化有微弱作用，锰离子和钙离子是抑制剂。5. 已发现二磷酸腺苷是一种竞争性抑制剂。6. 核糖体蛋白激酶催化反应的最大速度是每分钟每毫克蛋白有0.65纳摩尔的32P掺入氯化钾洗脱蛋白中。7. 以核糖体氯化钾洗脱蛋白为底物时的表观Km为0.71毫克/毫升，ATP的Km为94微摩尔。8. 通过在聚丙烯酰胺-十二烷基硫酸钠凝胶中电泳估计，核糖体蛋白激酶的分子量为60000，可能对应一个催化亚基。
