Cheneval D, Carafoli E
Laboratory of Biochemistry, Swiss Federal Institute of Technology, Zürich.
Eur J Biochem. 1988 Jan 15;171(1-2):1-9. doi: 10.1111/j.1432-1033.1988.tb13750.x.
It was recently shown that the mitochondrial isozyme of heart creatine kinase binds to cardiolipin on the outer half of the inner membrane [Müller, M., et al. (1985) J. Biol. Chem. 260, 3839-3843]. The enzyme has now been extracted and purified to homogeneity from rat heart mitochondria, and cleaved with CNBr. The fragments have been separated on an FPLC system using a Mono Q HR 5/5 column. Only one of these binds to cardiolipin-containing liposomes and has thus been identified as the cardiolipin-binding domain of the enzyme. Its amino acid sequence has been determined. The fragment contains 25 amino acids and corresponds to the N-terminal region of the protein. The binding of the fragment of cardiolipin-containing liposomes was inhibited by adriamycin. Another and larger CNBr fragment could be specifically labelled with periodate-oxidized (di-aldehyde) ATP and has thus been identified as the ATP-binding domain. Chemical modification of the basic amino acids Lys and Arg of the enzyme abolished its binding to cardiolipin.
最近有研究表明，心脏肌酸激酶的线粒体同工酶可与内膜外半部分的心磷脂结合[Müller, M., 等人 (1985) 《生物化学杂志》260, 3839 - 3843]。现已从大鼠心脏线粒体中提取并纯化该酶至同质状态，并用溴化氰进行切割。切割后的片段在配备Mono Q HR 5/5柱的快速蛋白质液相色谱（FPLC）系统上进行分离。其中只有一个片段能与含心磷脂的脂质体结合，因此被确定为该酶的心磷脂结合结构域。其氨基酸序列已被测定。该片段包含25个氨基酸，对应于该蛋白质的N端区域。阿霉素可抑制该心磷脂结合片段与含心磷脂脂质体的结合。另一个更大的溴化氰切割片段可以被高碘酸盐氧化（双醛）ATP特异性标记，因此被确定为ATP结合结构域。对该酶的碱性氨基酸赖氨酸和精氨酸进行化学修饰会消除其与心磷脂的结合。
