Heme Binding to Porphobilinogen Deaminase from Vibrio cholerae Decelerates the Formation of 1-Hydroxymethylbilane
Heme Binding to Porphobilinogen Deaminase from Vibrio cholerae Decelerates the Formation of 1-Hydroxymethylbilane
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DOI：
10.1021/acschembio.7b00934
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发表时间：
2018-03-01
影响因子：
4
通讯作者：
Ishimori, Koichiro
中科院分区：
文献类型：
--
作者：
Uchida, Takeshi;Funamizu, Takumi;Ishimori, Koichiro
胆色素原脱氨酶（PBGD）是一种在血红素生物合成过程中通过四个胆色素原分子的逐步聚合催化羟甲基胆烷（一种四吡咯中间体）形成的酶。在本研究中，来自霍乱弧菌的PBGD在大肠杆菌中表达并表征。出乎意料的是，光谱测量显示PBGD结合一当量的血红素，解离常数为0.33 +/- 0.01 μ M。吸收光谱和共振拉曼光谱表明，血红素是5配位和6配位血红素的混合物。突变研究表明，5-坐标血红素具有Cys 105作为血红素轴向配体，和His 227协调形成6-坐标血红素。血红素结合后，脱氨活性下降约15%。PBGD的晶体结构显示，His 227位于Cys 105附近，但His 227的侧链不指向Cys 105。此外，氰化物离子的血红素PBGD废除血红素结合酶活性的影响。因此，His 227与血红素的配位似乎诱导含有Cys 105的结构域的重定向，导致酶活性的降低。这是第一篇表明PBGD活性受血红素生物合成最终产物血红素控制的报道。这一发现提高了我们对血红素生物合成调节机制的理解。
Porphobilinogen deaminase (PBGD) is an enzyme that catalyzes the formation of hydroxymethylbilane, a tetrapyrrole intermediate, during heme biosynthesis through the stepwise polymerization of four molecules of porphobilinogen. PBGD from Vibrio cholerae was expressed in Escherichia coli and characterized in this study. Unexpectedly, spectroscopic measurements revealed that PBGD bound one equivalent of heme with a dissociation constant of 0.33 +/- 0.01 mu M. The absorption and resonance Raman spectra suggested that heme is a mixture of the 5-coordinate and 6-coordinate hemes. Mutational studies indicated that the 5-coordinate heme possessed Cys105 as a heme axial ligand, and His227 was coordinated to form the 6-coordinate heme. Upon heme binding, the deamination activity decreased by approximately 15%. The crystal structure of PBGD revealed that His227 was located near Cys105, but the side chain of His227 did not point toward Cys105. The addition of the cyanide ion to heme-PBGD abolished the effect of heme binding on the enzymatic activity. Therefore, coordination of His227 to heme appeared to induce reorientation of the domains containing Cys105, leading to a decrease in the enzymatic activity. This is the first report indicating that the PBGD activity is controlled by heme, the final product of heme biosynthesis. This finding improves our understanding of the mechanism by which heme biosynthesis is regulated.