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1.
Biochim Biophys Acta Proteins Proteom ; 1866(2): 348-355, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-29101077

RESUMO

Unexpected peptide deformylase (PDF) genes were recently retrieved in numerous marine phage genomes. While various hypotheses dealing with the occurrence of these intriguing sequences have been made, no further characterization and functional studies have been described thus far. In this study, we characterize the bacteriophage Vp16 PDF enzyme, as representative member of the newly identified C-terminally truncated viral PDFs. We show here that conditions classically used for bacterial PDFs lead to an enzyme exhibiting weak activity. Nonetheless, our integrated biophysical and biochemical approaches reveal specific effects of pH and metals on Vp16 PDF stability and activity. A novel purification protocol taking in account these data allowed strong improvement of Vp16 PDF specific activity to values similar to those of bacterial PDFs. We next show that Vp16 PDF is as sensitive to the natural inhibitor compound of PDFs, actinonin, as bacterial PDFs. Comparison of the 3D structures of Vp16 and E. coli PDFs bound to actinonin also reveals that both PDFs display identical substrate binding mode. We conclude that bacteriophage Vp16 PDF protein has functional peptide deformylase activity and we suggest that encoded phage PDFs might be important for viral fitness.


Assuntos
Amidoidrolases/química , Bacteriófagos/enzimologia , Vibrio parahaemolyticus/virologia , Proteínas Virais/química , Amidoidrolases/genética , Bacteriófagos/genética , Domínio Catalítico , Estabilidade Enzimática , Vibrio parahaemolyticus/genética , Proteínas Virais/genética
2.
Sci Rep ; 7(1): 11041, 2017 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-28887476

RESUMO

Prokaryotic proteins must be deformylated before the removal of their first methionine. Peptide deformylase (PDF) is indispensable and guarantees this mechanism. Recent metagenomics studies revealed new idiosyncratic PDF forms as the most abundant family of viral sequences. Little is known regarding these viral PDFs, including the capacity of the corresponding encoded proteins to ensure deformylase activity. We provide here the first evidence that viral PDFs, including the shortest PDF identified to date, Vp16 PDF, display deformylase activity in vivo, despite the absence of the key ribosome-interacting C-terminal region. Moreover, characterization of phage Vp16 PDF underscores unexpected structural and molecular features with the C-terminal Isoleucine residue significantly contributing to deformylase activity both in vitro and in vivo. This residue fully compensates for the absence of the usual long C-domain. Taken together, these data elucidate an unexpected mechanism of enzyme natural evolution and adaptation within viral sequences.


Assuntos
Amidoidrolases/química , Amidoidrolases/metabolismo , Bacteriófagos/enzimologia , Vibrio parahaemolyticus/virologia , Amidoidrolases/genética , Cristalografia por Raios X , Modelos Moleculares , Filogenia , Conformação Proteica
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