Oligomeric structure, enzyme kinetics, and substrate specificity of the phycocyanin alpha subunit phycocyanobilin lyase.
Oligomeric structure, enzyme kinetics, and substrate specificity of the phycocyanin alpha subunit phycocyanobilin lyase.
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DOI：
10.1016/s0021-9258(17)37022-9
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发表时间：
1994-03
期刊：
影响因子：
--
通讯作者：
C. Fairchild;A. Glazer
中科院分区：
文献类型：
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作者：
C. Fairchild;A. Glazer
藻胆蛋白携带线性四吡咯发色团（胆色素）硫醚连接到特定的半胱氨酸残基。在体内，胆色素与脱辅基蛋白的结合过程的特征在于，在一种藻胆蛋白上只有一个胆色素结合位点，即藻蓝蛋白的α亚基（α PC）上。在蓝细菌聚球藻属PCC 7002中，将藻蓝胆素添加至apo-α PC由cpcE和cpcF基因的蛋白产物催化。我们已经纯化并进一步表征了重组CpcE和CpcF蛋白。CpcE和CpcF形成酶活性1：1复合物（CpcEF），对尺寸排阻色谱稳定。CpcEF导致α PC荧光减少，并强烈影响其吸收光谱，但对β亚基没有影响。CpcEF胆色素添加活性相对于apo-α PC和胆色素表现出简单的Michaelis-Menten动力学。CpcEF还催化藻红胆素添加到apo-alpha PC;藻红胆素被认为是藻蓝胆素的生物合成途径。CpcEF显示藻蓝胆素相对于藻红胆素的偏好，无论是在结合亲和力和催化速率，足以说明藻蓝胆素选择性附着到apo-α PC。
Phycobiliproteins carry linear tetrapyrrole chromophores (bilins) thioether-linked to specific cysteine residues. The process of bilin attachment to apoprotein in vivo has been characterized for only one bilin attachment site on one phycobiliprotein, that on the alpha subunit of phycocyanin (alpha PC). In the cyanobacterium Synechococcus sp. PCC 7002, the addition of phycocyanobilin to apo-alpha PC is catalyzed by the protein products of the cpcE and cpcF genes. We have purified and further characterized the recombinant CpcE and CpcF proteins. CpcE and CpcF form an enzymatically active 1:1 complex (CpcEF), stable to size exclusion chromatography. CpcEF causes a reduction in alpha PC fluorescence and strongly affects its absorption spectrum but has no effect on the beta subunit. The CpcEF bilin addition activity exhibits simple Michaelis-Menten kinetics with respect to the apo-alpha PC and to bilin. CpcEF also catalyzes the addition of phycoerythrobilin to apo-alpha PC; phycoerythrobilin is thought to be on the biosynthetic pathway of phycocyanobilin. CpcEF shows a preference for phycocyanobilin relative to phycoerythrobilin, both in binding affinity and in the rate of catalysis, sufficient to account for selective attachment of phycocyanobilin to apo-alpha PC.