Meima Marcel E, Biondi Ricardo M, Schaap Pauline
School of Life Sciences, University of Dundee, Dundee DD1 5EH, United Kingdom.
Mol Biol Cell. 2002 Nov;13(11):3870-7. doi: 10.1091/mbc.e02-05-0285.
StmF mutants are chemotactic mutants that are defective in a cGMP phosphodiesterase (PDE) activity. We identified a novel gene, PdeD, that harbors two cyclic nucleotide-binding domains and a metallo-beta-lactamase homology domain. Similar to stmF mutants, pdeD-null mutants displayed extensively streaming aggregates, prolonged elevation of cGMP levels after chemotactic stimulation, and reduced cGMP-PDE activity. PdeD transcripts were lacking in stmF mutant NP377, indicating that this mutant carries a PdeD lesion. Expression of a PdeD-YFP fusion protein in pdeD-null cells restored the normal cGMP response and showed that PdeD resides in the cytosol. When purified by immunoprecipitation, the PdeD-YFP fusion protein displayed cGMP-PDE activity, which was retained in a truncated construct that contained only the metallo-beta-lactamase domain.
StmF突变体是在环鸟苷酸磷酸二酯酶(PDE)活性方面存在缺陷的趋化性突变体。我们鉴定出一个新基因PdeD,它含有两个环核苷酸结合结构域和一个金属β-内酰胺酶同源结构域。与stmF突变体相似,pdeD基因缺失突变体表现出广泛的流动聚集体,趋化刺激后cGMP水平的持续升高,以及cGMP-PDE活性降低。stmF突变体NP377中缺乏PdeD转录本,表明该突变体存在PdeD损伤。在pdeD基因缺失的细胞中表达PdeD-YFP融合蛋白可恢复正常的cGMP反应,并表明PdeD定位于细胞质中。通过免疫沉淀纯化时,PdeD-YFP融合蛋白表现出cGMP-PDE活性,该活性保留在仅包含金属β-内酰胺酶结构域的截短构建体中。