Formylation-independent eubacterial protein initiation

启动子甲硫酰-tRNA用于启动古细菌和真核生物细胞质的蛋白质合成。相反，真细菌和某些真核细胞器(如线粒体和叶绿体)的蛋白质合成可以使用启动子甲酰-甲硫酰-tRNA(fMet-tRNAfMet)启动。甲酰化是启动子甲硫酰-tRNA(Met-tRNAfMet)所特有的，由甲硫酰-tRNA甲酰基转移酶(MTF)催化。早期的体外和体内研究导致人们普遍认为，Met-tRNAfMet的甲酰化是启动所有真细菌蛋白质合成的关键步骤。但这一观点得到了加强，因为在大肠杆菌、肺炎链球菌和枯草杆菌中MTF基因的破坏严重抑制了细胞的生长，并且MTF基因存在于到目前为止测序的所有真细菌基因组中。然而，这种真细菌蛋白质启动的教条不再有效，因为一些细菌被证明启动蛋白质合成不依赖甲酰化。我的实验室第一个证明了真细菌铜绿假单胞菌具有利用Met-tRNAfMet启动蛋白质合成的能力。随后，研究证明，在特定的生长条件下，金黄色葡萄球菌、流感嗜血杆菌和酿酒酵母线粒体中Met-tRNAfMet的甲酰化对蛋白质的启动都不是必需的。因此，不同于依赖fMet-tRNAfMet启动蛋白质的大肠杆菌、肺炎链球菌和枯草杆菌，铜绿假单胞菌、金黄色葡萄球菌和流感嗜血杆菌都可以用fMet-tRNAfMet或Met-RNAfMet启动蛋白质。在这些细菌中，允许利用Met-tRNAfMet来启动蛋白质合成的原因还不清楚。此外，为什么一些细菌可以同时使用fMet-tRNAfMet和Met-tRNAfMet来启动蛋白质合成，而另一些细菌只使用fMet-tRNAfMet。我们研究的主要目的是(1)了解为什么一些细菌能够在不需要甲酰化的情况下启动蛋白质合成，而另一些细菌则依赖甲酰化来启动蛋白质合成，以及(2)鉴定其他不依赖甲酰化来启动蛋白质合成的真细菌。

The initiator methionyl-tRNA is used to initiate protein synthesis in archaebacteria and the cytoplasm of eukaryotes.  In contrast, protein synthesis in eubacteria and in certain eukaryotic organelles, such as mitochondria and chloroplasts, can be initiated using the initiator formyl-methionyl-tRNA (fMet-tRNAfMet).  Formylation is specific for the initiator methionyl-tRNA (Met-tRNAfMet) and is catalyzed by methionyl-tRNA formyltransferase (MTF). Early studies in vitro and in vivo led to the general belief that formylation of the Met-tRNAfMet is a key step in initiation of protein synthesis in all eubacteria.  This view was reinforced by the findings that disruption of the MTF gene in E. coli, S. pneumoniae and B. subtilis severely curtailed cell growth and the MTF gene is present in all eubacterial genomes sequenced to date.  However, this dogma of eubacterial protein initiation is no longer valid, since some bacteria have been shown to initiate protein synthesis independently of formylation. My laboratory was the first to demonstrate that a eubacteria, Pseudomonas aeruginosa, has the capacity to use Met-tRNAfMet to initiate protein synthesis.  Subsequently, it was demonstrated that formylation of Met-tRNAfMet was not necessary either for protein initiation in S. aureus, H. influenzae and Saccharomyces cerevisiae mitochondria under certain growth conditions.  Thus, unlike E. coli, S. pneumoniae and B. subtilis, which are dependent on fMet-tRNAfMet for protein initiation, P. aeruginosa, S. aureus, and H. influenzae can perform initiation with either fMet-tRNAfMet or Met-tRNAfMet.  The mechanism, however, that allows utilization of Met-tRNAfMet in protein initiation in these bacteria is not understood.  Furthermore, why some bacteria have the ability to initiate protein synthesis with both fMet-tRNAfMet and Met-tRNAfMet and others with only fMet-tRNAfMet is not understood.  The main goals of our research are to (1) understand why some bacteria have the ability to initiate protein synthesis without formylation while others rely on formylation, and (2) identify other eubacteria that are not dependent on formylation for protein initiation.