Muralytic activity of Micrococcus luteus Rpf and its relationship to physiological activity in promoting bacterial growth and resuscitation
Muralytic activity of Micrococcus luteus Rpf and its relationship to physiological activity in promoting bacterial growth and resuscitation
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DOI：
10.1111/j.1365-2958.2005.04930.x
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发表时间：
2006-01-01
影响因子：
3.6
通讯作者：
Young, M
中科院分区：
文献类型：
--
作者：
Mukamolova, GV;Murzin, AG;Young, M
几种放线菌的可培养性受复苏促进因子（Rpfs）控制。这些蛋白质含有一个c。70-残基结构域采用溶菌酶样折叠。在溶菌酶和各种细菌溶解性转糖基酶中发现的不变的催化谷氨酸残基在Rpf蛋白中也是保守的。来自藤黄微球菌（该蛋白质家族的创始成员）的Rpf确实是一种胞壁分解酶，正如含有M. luteus细胞壁和它的能力：（i）当表达和分泌到周质中时引起大肠杆菌裂解;（ii）从荧光胺标记的M.（iii）水解人工溶菌酶底物4-甲基伞形酮基-β-D-N，N ′，N ″-三乙酰壳三糖苷。当不变的谷氨酸残基被改变时，Rpf的活性降低，但没有完全消失。此外，在Rpf结构域中没有其他酸性残基是绝对需要的muralytic活性。替换一个或两个半胱氨酸残基，可能形成一个二硫桥内Rpf受损，但没有完全消除muralytic活性。Rpf突变体的胞壁溶解活性与其刺激细菌可培养性和复苏的能力相关，这与Rpf的生物活性直接或间接来自其裂解细菌肽聚糖中的键的能力的观点一致。
The culturability of several actinobacteria is controlled by resuscitation-promoting factors (Rpfs). These are proteins containing a c. 70-residue domain that adopts a lysozyme-like fold. The invariant catalytic glutamate residue found in lysozyme and various bacterial lytic transglycosylases is also conserved in the Rpf proteins. Rpf from Micrococcus luteus, the founder member of this protein family, is indeed a muralytic enzyme, as revealed by its activity in zymograms containing M. luteus cell walls and its ability to (i) cause lysis of Escherichia coli when expressed and secreted into the periplasm; (ii) release fluorescent material from fluorescamine-labelled cell walls of M. luteus; and (iii) hydrolyse the artificial lysozyme substrate, 4-methylumbelliferyl-beta-D-N,N',N''-triacetylchitotrioside. Rpf activity was reduced but not completely abolished when the invariant glutamate residue was altered. Moreover, none of the other acidic residues in the Rpf domain was absolutely required for muralytic activity. Replacement of one or both of the cysteine residues that probably form a disulphide bridge within Rpf impaired but did not completely abolish muralytic activity. The muralytic activities of the Rpf mutants were correlated with their abilities to stimulate bacterial culturability and resuscitation, consistent with the view that the biological activity of Rpf results directly or indirectly from its ability to cleave bonds in bacterial peptidoglycan.