SUBUNIT LOCATION AND SEQUENCES OF THE CYSTEINYL PEPTIDES OF PIG-HEART NAD-DEPENDENT ISOCITRATE DEHYDROGENASE
SUBUNIT LOCATION AND SEQUENCES OF THE CYSTEINYL PEPTIDES OF PIG-HEART NAD-DEPENDENT ISOCITRATE DEHYDROGENASE
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DOI：
10.1021/bi00488a010
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发表时间：
1990-09-11
期刊：
影响因子：
2.9
通讯作者：
COLMAN, RF
中科院分区：
文献类型：
--
作者：
HUANG, YC;COLMAN, RF
猪心NAD依赖性异柠檬酸脱氢酶具有由α 2 β组成的亚基结构。γ，用α。分子量为39000的β亚基和分子量为39000的β亚基。和γ每一种的分子量为41000。氨基末端序列（33-35个残基）和半胱氨酰肽序列现在已经通过使用由色谱聚焦或等电聚焦和电印迹分离的亚基来确定。α的N-末端序列的置换。β亚基的氨基酸残基相对于较大β亚基的氨基酸残基减少11-12个氨基酸。和γ亚基揭示了17个氨基酸区域的高度相似性，其中10个残基在所有三个亚基中相同。完整的酶有6.0个游离SH基团，平均亚单位为40000道尔顿，但产生15个可区分的半胱氨酸在分离的胰蛋白酶肽。测序肽中的六个不同的半胱氨酸已经定位于α-半胱氨酸。亚单位的β和γβ亚基分别含有7个和5个半胱氨酸，含有3个半胱氨酸的胰蛋白酶肽是β亚基共有的。和γ亚单位。这三个亚基似乎密切相关，但β亚基与β亚基之间的关系并不密切。和γ彼此之间的相似性大于与α的相似性。亚单位来自猪心的NAD特异性异柠檬酸脱氢酶已显示具有2个异柠檬酸、锰离子、NAD+和变构激活剂ADP的结合位点/酶四聚体[Colman，R. F. 04 The Dog（1983）Protein Rev.1，41-49]。提出催化活性的四聚体酶被组织为二聚体的二聚体，其中α。β的和α。γ的二聚体是不同的，但功能相似。
Pig heart NAD-dependent isocitrate dehydrogenase has a subunit strucutre consisting of .alpha.2.beta..gamma., with the .alpha. subunit exhibitng a molecular weight of 39000 and the .beta. and .gamma. each having molecular weights of 41000. The amino-terminal sequences (33-35 residues) and the cysteinyl peptide sequences have now been determined by using subunits separated by chromatofocusing or isoelectric focusing and electroblotting. Displacement of the N-terminal sequence of the .alpha. subunit by 11-12 amino acids relative to that of the larger .beta. and .gamma. subunits reveals a 17 amino acid region of great similarity in which 10 residues are identical in all three subunits. The complete enzyme has 6.0 free SH groups per average subinit of 40000 daltons, but yields 15 distinguishable cysteines in isolated tryptic peptides. Six distinct cysteines in sequenced peptides have been located in the .alpha. subunit. The .beta. and .gamma. subunits contain seven and five cysteines, respectively, with tryptic peptide containing three cysteines being common to the .beta. and .gamma. subunits. The three subunits appear to be closely related, but .beta. and .gamma. are similar to each other than either is to the .alpha. subunit. The NAD-specific isocitrate dehydrogenase from pig heart has been shown to have 2 binding sites/enzyme tetramer for iscitrate, manganous ion, NAD+, and allosteric activator ADP [Colman, R. F. (1983) Pept. Protein Rev. 1, 41-49]. It is proposed that the catalytically active tetrameric enzyme is organized as a dimer of dimers in which the .alpha..beta. and .alpha..gamma. dimers are nonidentical but functionally similar.