N2和NH4^＋培养下粪产碱菌固氮酶铁蛋白圆二色谱及热力学特性比较

The CD Spectra and Thermodynamic Properties of the Iron Protein of Nitrogenase from N_2-and NH_4￣+ -Grown Alcaligenes faecalis

Ｎ２和ＮＨ培养下粪产碱菌固氮酶铁蛋白（分别为Ａｆ２和Ａｆ＊２）的氧化态和还原态的ＣＤ谱及ＭＣＤ谱存在明显差异。还原态Ａｆ２与Ａｆ＊２在２１０ｎｍ附近的ＭＣＤ谱完全不同，但它们的氧化态ＭＣＤ谱相同。热力学测定结果表明．在２９８°Ｋ，１．０１３×１０５Ｐａ下，Ａｆ２与ＭｇＡＴＰ饱合络合时的培（△Ｈ°）变化为－２７．０ｋＪ／ｍｏｌ，自由能（△Ｇ°）变化则为－２０．８ｋＪ／ｍｏｌ．熵（△Ｓ°）变４也为－２０．６Ｊｍｏｌ－１Ｋ－１。而Ａｆ＊２与ＭｇＡＴＰ饱合络合时的烂变化为－４４．３５ｋＪ／ｍｏｌ，自由能变化为一２０．４ｋＪ／ｍｏｌ，熵变化为－７９．９Ｊｍｏｌ－１Ｋ－１。Ａｆ２与ＭｇＡＴＰ络合后再与Ｎ２培养的钼铁蛋白反应为吸热反应，而Ａｆ＊２与ＭｇＡＴＰ络合后再与ＮＨ培养的钼铁蛋白反应为放热反应。

The purified iron proteins (Af2 and Af* 2) of nitrogenase from Alcalgigenes faecalis strain A1501 cells grown under N2 and under NH (30 mmol/ L) respectively were used in this study. The specific activity of Af2 was 1540 nmol C2H4 formed mg-1 protein min-1, and no activity of Af* 2 was observed. The circular dichoism (CD) and magnetic CD (MCD) were measured in both reduced and oxidized states. Af2 and Af* 2 differed from each other in their CD spectra, particularly in the oxidized saltes. After oxidization, the 210 urn ark of the CDspectra from Af2 was shifted to the far-UV region, while the spectra of Af*2 was almOSt unchanged. The MCD spectra of the reduced states of Af2 and Af*2 were quite different, but these of the oxidized saltes were the same, suggesting that their molecular structure and the environmental conditions around the 4Fe-4S cluster might be different.The binding of MgATP to Af2 and Af* 2 has been studied using isothermal Calorimeter, and a △H° value of-27. 0 kJ/mol for Af2 and 44. 35 kJ/mol for Af*2 were measured at 298 °K under 1. 013×105Pa. Such enthalpy values combinedwith a △G° value of-20. 8 kJ/mol for Af2 and -20. 4 kJ/mol for Af* 2cave fis' values of- 20. 6 J mol-1K-1 for Af2 and - 79. 9 J mol-1 K-1for Af* 2. respectively. The K. forAf2 and Af* 2 bound MgATP were 222 mmol/L and 263 mmol/L, respectively. The Af2. MgATP and Af*2. MgATP were titrated with Afl and Af*1,and the △H° value for Af2. MgATP-Afl was + 29. 8 kJ/moland that for Af*2. MgATP-Af*1was-60. 2 kJ/mol. The former wasan endothermic reaction, while the latter was exothermic one. The thermodynamic parameters also suggested that molecular structure of Af2 and Af* 2 might be different.