Howard W D, Solomonson L P
J Biol Chem. 1981 Dec 25;256(24):12725-30.
Chlorella nitrate reductase catalyzes the reduction of nitrate to nitrite by NADH. Initial velocity studies showed that the kinetic mechanism is sequential, indicating that both substrates must bind to the enzyme before any products are released. Product inhibition with NAD and nitrite showed that competitive inhibition was observed when the inhibitor was similar to the varied substrate, while noncompetitive inhibition was observed when the inhibitor was dissimilar to the varied substrate. Likewise, dead-end inhibition with adenosine 5'-diphosphoribose and thiocyanate showed competitive inhibition when the inhibitor was similar to the varied substrate and noncompetitive inhibition when the inhibitor was dissimilar to the varied substrate. These results indicate that Chlorella nitrate reductase follows a random bi bi kinetic mechanism. Phosphate was found to stimulate NADH:nitrate reductase activity and 2-fold. The NADH:cytochrome c reductase activity associated with nitrate reductase was not affected by phosphate suggesting the effect of phosphate is on the nitrate-reducing moiety of the enzyme. Phosphate increases Vmax but has no effect on the apparent Km for nitrate.
小球藻硝酸还原酶催化由NADH将硝酸盐还原为亚硝酸盐。初始速度研究表明，动力学机制是有序的，这表明在任何产物释放之前，两种底物都必须与酶结合。用NAD和亚硝酸盐进行的产物抑制表明，当抑制剂与可变底物相似时，观察到竞争性抑制，而当抑制剂与可变底物不同时，观察到非竞争性抑制。同样，用腺苷5'-二磷酸核糖和硫氰酸盐进行的终产物抑制表明，当抑制剂与可变底物相似时，观察到竞争性抑制，而当抑制剂与可变底物不同时，观察到非竞争性抑制。这些结果表明，小球藻硝酸还原酶遵循随机双底物双产物动力学机制。发现磷酸盐可刺激NADH：硝酸还原酶活性达2倍。与硝酸还原酶相关的NADH：细胞色素c还原酶活性不受磷酸盐影响，这表明磷酸盐的作用是针对酶的硝酸盐还原部分。磷酸盐增加Vmax，但对硝酸盐的表观Km没有影响。
