STUDIES ON FTIR SPECTRA OF SOLUBLE HYDROGENASE FROM CHROMATIUM VINOSUM

光合细菌Chromatiumvinosum含有一种可溶性氢酶和一种膜结合态氢酶。氧化态可溶性氢酶在红外光谱区(1860 -2140cm -1)有四个特征吸收峰 (2103.7 ,2086.2,2054.8和1962.5cm -1)。其中1962.5cm -1处吸收带的位置与已知的NiFe -氢酶活性中心 -CO基团所产生的吸收带位置相近 ;另外三条吸收带的位置与已知的NiFe -氢酶活性中心 -CN基团所产生的吸收带的位置相近。以2 ,6 -二氯酚靛酚 (DPIP)氧化可溶性氢酶时 ,四条吸收谱带的位置基本上没有发生变化。可溶性氢酶被Na2S2O4 充分还原时,-CO基团的吸收带移至1946.8cm -1 ,而 -CN基团的三条吸收带中的两条分别移至2076.8cm-1 和2093.1cm-1 处 ,另一条则消失了。还原态可溶性氢酶与CO反应后 ,其红外光谱显示七条吸收带 ,在 -CO基团红外光谱区和 -CN基团红外光谱区各产生了两条新的吸收带。研究表明 ,C.vinosum可溶性氢酶的活性中心的结构类似于其它已知的NiFe -氢酶 ,但与活性中心金属原子相连的可能包括三个 -CN基团和一个 -CO基团 ,结合可溶性氢酶的EPR谱特征 ,推测C.vinosum可溶性氢酶活性中心的结构可能为Ni(CN)Fe(CN)2(CO)。Fourier transform infrared (FTIR) studies of oxidized soluble hydrogenase (SH) from C.vinosum revealed four absorption bands (2103.7, 2086.2, 2054.8 and 1962.5cm-1) in the 1860-2140cm-1 region. A strong band at 1962.5cm-1 is in the same region as the band due to the intrinsic -CO group bound to Fe in the active site of other NiFe-hydrogenases. The other three bands(2054.8, 2086.3 and 2103.7cm-1) are similar in position to those arising from the stretch vibrations of -CN group in other NiFe-hydrogenase. When the soluble hydrogenase was completely oxidized by 2,6-dichlorophenolindophenol, no band shift was observed. When the soluble hydrogenase was fully reduced by Na2S2O4, all the bands shifted. The ν(CO) band shifted to 1946.8cm-1.The other bands moved to 2076.8cm-1 and 2093.1cm-1. Addition of CO to the reduced enzyme resulted in changes of both the number and position of the bands. Two new bands (1954.7cm-1 and 1967.7cm-1) were observed in the -CO region, while two new bands (2063.9cm-1 and 2071.7cm-1) showed up in the -CN region.教育部重点科技项目资助!(No:99070