Biswas S B, Biswas E E
J Biol Chem. 1987 Jun 5;262(16):7831-8.
The dnaB protein of Escherichia coli, a multifunctional DNA-dependent ribonucleotide triphosphatase and dATPase, cross-links to ATP on ultraviolet irradiation under conditions that support rNTPase and dATPase activities of dnaB protein. The covalent cross-linking to ATP is specifically inhibited by ribonucleotides and dATP. Tryptic peptide mapping demonstrates that ATP cross-links to only the 33-kDa tryptic fragment (Fragment II) of dnaB protein. The presence of single-stranded DNA alters the covalent labeling of dnaB protein by ATP, suggesting a possible role of DNA on the mode of nucleotide binding by dnaB protein. Present studies demonstrate that the dnaC gene product binds ribonucleotides independent of dnaB protein. On dnaB-dnaC protein complex formation, covalent incorporation of ATP to dnaB protein decreases approximately 70% with a concomitant increase of ATP incorporation to dnaC protein by approximately 3-fold. The mechanism of this phenomenon has been analyzed in detail by titrating dnaB protein with increasing amounts of dnaC protein. The binding of dnaC protein to dnaB protein appears to be a noncooperative process. The lambda P protein, which interacts with dnaB protein in the bacteriophage lambda DNA replication, does not bind ATP in the presence or absence of dnaB protein. However, lambda P protein enhances the covalent incorporation of ATP to dnaB protein approximately 4-fold, suggesting a direct physical interaction between lambda P and dnaB proteins with a probable change in the modes of nucleotide binding to dnaB protein. The lambda P protein likely forms a lambda P-dnaB-ATP dead-end ternary complex. The implications of these results in the E. coli and bacteriophage lambda chromosomal DNA replication are discussed.
大肠杆菌的DnaB蛋白是一种多功能的依赖DNA的核糖核苷酸三磷酸酶和dATP酶，在支持DnaB蛋白的rNTPase和dATPase活性的条件下，紫外线照射时它会与ATP发生交联。与ATP的共价交联受到核糖核苷酸和dATP的特异性抑制。胰蛋白酶肽图谱分析表明，ATP仅与DnaB蛋白的33 kDa胰蛋白酶片段（片段II）发生交联。单链DNA的存在改变了ATP对DnaB蛋白的共价标记，这表明DNA可能在DnaB蛋白结合核苷酸的模式中发挥作用。目前的研究表明，DnaC基因产物独立于DnaB蛋白结合核糖核苷酸。在形成DnaB-DnaC蛋白复合物时，ATP与DnaB蛋白的共价掺入减少约70%，同时ATP与DnaC蛋白的掺入增加约3倍。通过用越来越多的DnaC蛋白滴定DnaB蛋白，对这一现象的机制进行了详细分析。DnaC蛋白与DnaB蛋白的结合似乎是一个非协同过程。在噬菌体λ DNA复制中与DnaB蛋白相互作用的λ P蛋白，无论有无DnaB蛋白都不结合ATP。然而，λ P蛋白使ATP与DnaB蛋白的共价掺入增加约4倍，这表明λ P蛋白与DnaB蛋白之间存在直接的物理相互作用，并且核苷酸与DnaB蛋白结合模式可能发生变化。λ P蛋白可能形成λ P-DnaB-ATP终产物三元复合物。讨论了这些结果对大肠杆菌和噬菌体λ染色体DNA复制的影响。
