Discovery of a New Genetic Variant of Methionine Aminopeptidase from Streptococci with Possible Post-Translational Modifications: Biochemical and Structural Characterization

Abstract

Protein N-terminal methionine excision is an essential co-translational process that occurs in the cytoplasm of all organisms. About 60-70% of the newly synthesized proteins undergo this modification. Enzyme responsible for the removal of initiator methionine is methionine aminopeptidase (MetAP), which is a dinuclear metalloprotease. This protein is conserved through all forms of life from bacteria to human except viruses. MetAP is classified into two isoforms, Type I and II. Removal of the map gene or chemical inhibition is lethal to bacteria and to human cell lines, suggesting that MetAP could be a good drug target. In the present study we describe the discovery of a new genetic variant of the Type I MetAP that is present predominantly in the streptococci bacteria. There are two inserts (insert one: 27 amino acids and insert two: four residues) within the catalytic domain. Possible glycosylation and phosphorylation posttranslational modification sites are identified in the ‘insert one’. Biochemical characterization suggests that this enzyme behaves similar to other MetAPs in terms of substrate specificity. Crystal structure Type Ia MetAP from Streptococcus pneumoniae (SpMetAP1a) revealed that it contains two molecules in the asymmetric unit and well ordered inserts with structural features that corroborate the possible posttranslational modification. Both the new inserts found in the SpMetAP1a structurally align with the P-X-X-P motif found in the M. tuberculosis and human Type I MetAPs as well as the 60 amino acid insert in the human Type II enzyme suggesting possible common function. In addition, one of the β-hairpins within in the catalytic domain undergoes a flip placing a residue which is essential for enzyme activity away from the active site and the β-hairpin loop of this secondary structure in the active site obstructing substrate binding. This is the first example of a MetAP crystallizing in the inactive form.

A mutation in the methionine aminopeptidase gene provides phage resistance in Streptococcus thermophilus

“…While the MetAP of S . thermophilus has not been previously studied, it shares 89% identity and 96% similarity with the MetAP of Streptococcus pneumoniae TIGR4 for which the structure has been determined (PDB 4KM3) 44 . Although the histidine at position 206 was not annotated as part of the active site (Fig.…”

Catabolite control protein A is an important regulator of metabolism in Streptococcus suis type 2

“…These, and other genes, were found to have extremely important effects for the methionine acquisition and synthesis of Streptococcus pneumoniae in the growth and virulence when using gene deletions methods (24). By contrast, the methionine aminopeptidase is a dinuclear metalloprotease, which is conserved in all forms of life in bacteria (25). Others, including the glutamine, glutamate and uridine biosynthesis and transport, are stringently controlled during bacteria metabolism (26)(27)(28).…”

Comparing extracellular proteins from thermostable direct haemolysin‐related haemolysin‐positive and ‐negativeVibrio parahaemolyticusstrains to identify potential virulence factors

“…Methionine aminopeptidase (vps17802‐4) is responsible for removing the initial methionine during protein processing and plays a pivotal role in cell growth (Gonzales & Robert‐Baudouy, 1996). In some bacteria, such as Acinetobacter baumannii , E. coli and Streptococci , it promotes their adaptability to the environment and also contributes to virulence (Arya et al., 2013; Chai et al., 2008; Yuan et al., 2011).…”