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本文报道半胱氨酸、赖氨酸和精氨酸等残基在鸡肝果糖-6-磷酸-2-激酶的作用。DTNB修饰的结果表明鸡肝PFK-2每亚基具有9个巯基。其中,在中性pH下可滴定的巯基数目为6个;在Fru6P存在下的可滴定巯基数为4个;而在ATP存在下的可滴定巯基数为7个。在修饰的过程中酶的活力随着被滴定的巯基数目的增加先是提高后为下降,但最后的酶仍不低于初始活力。NEMI对该酶活力的影响与DTNB的相似,它的修饰首先使酶活力升高,当巯基完全被修饰后的活力仍不低于初始活力。但是PCMB和碘代乙酸对该酶的活力影响很小。pH动力学结果显示该酶有1个pK~a为9.0的必需基团,它很可能是赖氨酸残基。PLP和苯乙二醛修饰PFK-2皆可导致该酶的失活。Fru6P对PLP的作用有强烈的保护,Fru2,6P2和Pi也有一定的保护作用,而ATP则稍有增加PLP对该酶的失活作用;苯乙二醛修饰的结果和PLP不同,Fru2,6P2有较强的保护作用,ATP有部分保护作用,Fru6P则对失活完全无保护作用。以上结果表明赖氨酸残基是鸡肝PFK-2与底物Fru2,6P2结合有关的必需残基,而精氨酸可能是该酶与产物Fru2,6P2结合有关的残基。
This paper reports the effects of residues of cysteine, lysine and arginine on hen-fructose-6-phosphate-2-kinase. The result of DTNB modification shows that there are 9 mercapto groups per subunit of chicken liver PFK-2. Among them, the number of titratable thiol groups at neutral pH was 6; the number of titratable thiol groups in the presence of Fru6P was 4; and the number of titratable thiol groups in the presence of ATP was seven. In the process of modification, the enzyme activity first increases and then decreases with the increase of titration thiol number, but the final enzyme is still not lower than the initial activity. The effect of NEMI on the enzyme activity is similar to that of DTNB. Its modification firstly increases the enzyme activity, and the activity after the thiol group is completely modified is still not lower than the initial activity. However, PCMB and iodoacetic acid have little effect on the enzyme activity. The pH kinetic results show that the enzyme has one essential group with pK ~ a of 9.0, which is likely to be a lysine residue. PLP and phenylglyoxal modified PFK-2 can lead to the enzyme inactivation. Fru6P has a strong protective effect on PLP, Fru2,6P2 and Pi also have a protective effect, while ATP slightly increased PLP on the enzyme inactivation; phenylglyoxal modified results and PLP is different, Fru2, 6P2 There is a strong protective effect, ATP has some protective effect, Fru6P completely devoid of protective effect. The above results indicate that the lysine residue is an essential residue related to the binding of the chicken liver PFK-2 to the substrate Fru2,6P2, and arginine may be the residue of the enzyme that binds to the product Fru2,6P2.