酵母羧甲基化及光照甘油醛-3-磷酸脱氢酶在碘化钾溶液中荧光发色团的形成

CM-GAPDH在碘化钾溶液中,NAD~+的存在下,形成发射波长为383nm的荧光物。对照的NAD~+与碘化钾溶液混合不产生荧光物。全位及半位修饰光照酶的内源荧光在碘化钾溶液中的变化与天然酶的有明显不同。两者在碘化钾中都形成383nm的荧光,但全位修饰光照酶形成383nm荧光的最适碘化钾浓度为1.0M;半位修饰的为0.8M。以上结果暗示:383nm荧光物的形成需要GAPDH和NAD~+同时存在,并且与活性部位巯基修饰的多少有关,该荧光物可能位于GAPDH的活性部位。

FORMATION OF POTASSIUM IODIDE FLUORESCENT DERIVATIVES OF CARBOXYMETHYLATION AND NAD DERIVATIVES OFYEAST D-GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE

Although tetrakiscarboxymethylated or biscarboxymethylated enzyme has not formed the fluorophore emitting 383 nm fluorescence with potassium iodide, each of them can emit the fluorescence after incubated with NAD+ for 30 h in the solution, but the control sample has little appreciable 383 nm emission by NAD+ dissolved with the potassium iodide. Nevertheless, NAD fluorescence derivative corresponding the tetrakiscarboxymethylated and biscarboxy-thylated enzyme can form the fluorophore of 383 nm with potassium iodide. Some differences in concentration of potassium iodide for optimum formation of 383 nm fluorophore between the tetrakiscarboxymethylated and the biscar-boxymethylated enzymes, as well as their corresponding NAD derivatives have been demonstrated. It appears that the fluorophore derivative may be near or at the active site, since formation of 383 nm fluorescence demands of GAPDH together with NAD + , which is appreciably effected by the amount of modific-cation of Cys-149.