Modulation of hyperthermophilic DNA polymerase activity by archaeal chromatin proteins
Modulation of hyperthermophilic DNA polymerase activity by archaeal chromatin proteins
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DOI：
10.1074/jbc.m309860200
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发表时间：
2004-01-02
影响因子：
4.8
通讯作者：
Huang, L
中科院分区：
文献类型：
--
作者：
Lou, HQ;Duan, ZH;Huang, L
Sulfolobus合成大量高度保守的7-kDa dna结合蛋白，怀疑与染色体组织有关。研究了7-kDa蛋白对嗜热古细菌solfataricus Sulfolobus solfataricus B族DNA聚合酶（polB1）聚合和3′-5′外切酶活性的影响。polB1在生理相关的温度下以相似的速率降解单链DNA和双链DNA。7-kDa蛋白能够显著抑制聚合酶对匹配模板引物的切除并增强其延伸。然而，这些蛋白不能保护单链DNA免受polB1的切割。此外，7-kDa蛋白不影响polB1的校对能力，也不抑制聚合酶对不匹配引物的切除。有效抑制polB1的3‘-5’外切酶活性所需的dNTP浓度从缺乏7-kDa蛋白时的约1 mM降低到65℃下存在蛋白质时的约50 mM。我们的数据表明，7-kDa染色质蛋白可以调节超嗜热DNA聚合酶的延伸和切除活性，降低酶在高温下的校对成本。
Sulfolobus synthesizes a large quantity of highly conserved 7-kDa DNA-binding proteins suspected to be involved in chromosomal organization. The effect of the 7-kDa proteins on the polymerization and 3'-5' exonuclease activities of a family B DNA polymerase ( polB1) from the hyperthermophilic archaeon Sulfolobus solfataricus was investigated. polB1 degraded both single-stranded DNA and double-stranded DNA at similar rates in vitro at temperatures of physiological relevance. The 7-kDa proteins were capable of significantly inhibiting the excision and enhancing the extension of matched template primers by the polymerase. However, the proteins did not protect single-stranded DNA from cleavage by polB1. In addition, the 7-kDa proteins did not affect the proofreading ability of polB1 and were not inhibitory to the excision of mismatched primers by the polymerase. The dNTP concentrations required for the effective inhibition of the 3'-5' exonuclease activity of polB1 were lowered from similar to1 mM in the absence of the 7-kDa proteins to similar to50 muM in the presence of the proteins at 65 degreesC. Our data suggest that the 7-kDa chromatin proteins serve to modulate the extension and excision activities of the hyperthermophilic DNA polymerase, reducing the cost of proofreading by the enzyme at high temperature.