Ca^(2+)和Mg^(2+)对野生型和变异型牙龈卟啉单胞菌FtsZs的GTPase活性的影响

Effects of Ca^(2+) and Mg^(2+) on GTPase activities of wild-type and mutant FtsZs in Porphyromonas gingivalis

目的:探讨Ca2+和Mg2+对野生型和变异型牙龈卟啉单胞菌FtsZs的GTPase活性的影响。方法:将质粒pEZ1(野生型PgFtsZ,Wt PgFtsZ)、pYW1(PgFtsZ的C末端缺失73个氨基酸残基的变异型,ZΔC01)和pYW2(PgFtsZ的N末端缺失43个氨基酸残基的变异型,ZΔN01)导入大肠杆菌BL21(DE3)pLysS中表达,分离和纯化这些目的蛋白。应用Malachite green检测法检测10mmol.L-1的Ca2+、Mg2+对Wt PgFtsZ、ZΔC01和ZΔN01的GTPase活性的作用。结果:在没有10mmol.L-1Ca2+或Mg2+的条件下,Wt PgFtsZ、ZΔC01和ZΔN01的GTPase值分别为8.25±0.22、7.43±0.23和8.00±0.18;在10mmol.L-1Mg2+存在的条件下,Wt PgFtsZ、ZΔC01和ZΔN01的GTPase值分别为8.30±0.15、7.51±0.22和7.85±0.17,同没有离子存在的条件下相比GTPase活性差异均无显著性(P>0.05);在10mmol.L-1Ca2+存在的条件下,Wt PgFtsZ、ZΔC01和ZΔN01的GTPase值分别为5.84±0.20、5.25±0.18和5.73±0.13,同没有离子存在的条件下比较GTPase活性均有明显降低(P<0.01)。结论:10mmol.L-1的Mg2+对Wt PgFtsZ、ZΔC01和ZΔN01的GTPase活性无影响,10mmol.L-1的Ca2+则对GTPase具有抑制作用。

Objective To investigate the effects of Ca^2＋ and Mg^2＋ on the GTPase activities of the wild-type and mutant FtsZs in Porphyromonas gingivalis. Methods Plasmids pEZ1 （Wt PgFtsZ）, pYW1 （missing 73 residues from the C-terminus of PgFtsZ, ZAC01） and pYW2 （missing 43 residues from the N-terminus of PgFtsZ, ZAN01） were introduced into Escherichia coli BL21 （DE3） pLysS to express and purify the purpose proteins. Malachite green assay was used to measure the GTPase activities of purified Wt PgFtsZ, ZΔCO1 and ZΔNO1. Results GTPase activities of Wt PgFtsZ, ZΔC01 and ZΔN01 were 8.25±0.22, 7.43±0.23, 8.00±0.18, respectively, without no ion. GTPase activities of Wt PgFtsZ, ZΔC01 and ZΔN01 were 8.30 -± 0.15, 7.51 ± 0.22, 7.85 ± 0.17, respectively, with 10 mmol · L^-1 Mg^2＋ , and not affected by 10 mmol · L^-1 Mg^2＋（P〉 0.05） . However, GTPase activities of Wt PgFtsZ, ZΔC01 and ZΔN01 were 5.84±0.20, 5.25±0.18, 5.73±0.13, respectively, with 10 mmol ·L^-1 Ca^2＋ , and reduced by 10 mmol ·L^-1 Ca^2＋（P〈0.01） . Conclusion 10 mmol ·L^-1 Mg^2＋ has no effect on GTPase activities of Wt PgFtsZ, ZΔC01 and ZΔN01. However, 10 mmol ·L^-1 Ca^2＋ inhibits their GTPase activities.