灵芝中一种新的脱氧核糖核酸酶的纯化及特征

通过(NH4)2SO4沉淀,利用Blue Sepharose 6 Fast Flow和SP Sepharose Fast Flow层析方法从灵芝中分离纯化了一种脱氧核糖核酸酶,命名为GLDNase.通过质谱分析确定GLDNase精确分子质量为13807 u.SDS-PAGE电泳表明GLDNase为单亚基多肽,其N-端氨基酸序列为PLDTGRYHIYTW/T/CDGG.GLDNase可作用于ssDNA和dsDNA,是一种非限制性内切酶,酶活性依赖于二价金属阳离子Mg2+,10 mmol.L-1EDTA可完全抑制其活性.最适pH值为8.4,40℃时相对活性最高.GLDNase水解DNA的产物末端的基团为3′-OH、5′-磷酸.

Purification and characterization of a novel deoxyriboendonuclease from Ganoderma lucidum

A novel deoxyribonuclease from the fruiting bodies of Ganoderma lucidum has been purified and designated as GLDNase.It was purified by ammonium sulphate precipitation,Blue Sepharose 6 Fast Flow and SP Sepharose Fast Flow.GLDNase showed a single protein band on SDS-PAGE.It showed a major mass peak of 13807 u analyzed in an electrospray-mass analyzer.Amino acid analysis of the N-terminus indicated that the sequence was PLDTGRYHIYTW/T/CDGG.GLDNase acted on both ssDNA and dsDNA.Its optimum pH and temperature were 8.4 and 40 ℃ respectively.It was a kind of non-restriction-endonuclease.Mg2+ was required for the endonuclase activity as a co-factor.The activity of the purified enzyme was completely inhibited by 10 mmol·L-1 EDTA.The oligonucleotides produced by GLDNase had a phosphate group at the 5'-termini and a hydroxyl group at the 3'-termini.