Structure and function of a serine carboxypeptidase adapted for degradation of the protein synthesis antibiotic microcin C7

被引:28
作者
Agarwal, Vinayak [1 ,2 ]
Tikhonov, Anton [4 ,5 ,7 ]
Metlitskaya, Anastasia [4 ,5 ]
Severinov, Konstantin [4 ,5 ,6 ,7 ]
Nair, Satish K. [1 ,2 ,3 ]
机构
[1] Univ Illinois, Ctr Biophys & Computat Biol, Urbana, IL 61801 USA
[2] Univ Illinois, Inst Genom Biol, Urbana, IL 61801 USA
[3] Univ Illinois, Dept Biochem, Urbana, IL 61801 USA
[4] Russian Acad Sci, Inst Mol Genet, Moscow 11934, Russia
[5] Russian Acad Sci, Inst Gene Biol, Moscow 11934, Russia
[6] Rutgers State Univ, Dept Mol Biol & Biochem, Piscataway, NJ 08854 USA
[7] Rutgers State Univ, Waksman Inst, Piscataway, NJ 08854 USA
基金
俄罗斯基础研究基金会;
关键词
tRNA synthetase inhibitor; self-immunity; reaction mechanism; protein engineering; TRANSFER-RNA SYNTHETASE; ALPHA-CHYMOTRYPSIN; HYDROLYSIS; PEPTIDASE; MECHANISM; SPECIFICITY; PROTEASES; TRIAD; BOND;
D O I
10.1073/pnas.1114224109
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
Several classes of naturally occurring antimicrobials exert their antibiotic activity by specifically targeting aminoacyl-tRNA synthetases, validating these enzymes as drug targets. The aspartyl tRNA synthetase "Trojan horse" inhibitor microcin C7 (McC7) consists of a nonhydrolyzable aspartyl-adenylate conjugated to a hexapeptide carrier that facilitates active import into bacterial cells through an oligopeptide transport system. Subsequent proteolytic processing releases the toxic compound inside the cell. Producing strains of McC7 must protect themselves against autotoxicity that may result from premature processing. The mccF gene confers resistance against endogenous and exogenous McC7 by hydrolyzing the amide bond that connects the peptide and nucleotide moieties of McC7. We present here crystal structures of MccF, in complex with various ligands. The MccF structure is similar to that of dipeptide LD-carboxypeptidase, but with an additional loop proximal to the active site that serves as the primary determinant for recognition of adenylated substrates. Wild-type MccF only hydrolyzes the naturally occurring aspartyl phosphoramidate McC7 and synthetic peptidyl sulfamoyl adenylates that contain anionic side chains. We show that substitutions of two active site MccF residues result in a specificity switch toward aromatic aminoacyl-adenylate substrates. These results suggest how MccF-like enzymes may be used to avert various toxic aminoacyl-adenylates that accumulate during antibiotic biosynthesis or in normal metabolism of the cell.
引用
收藏
页码:4425 / 4430
页数:6
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