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Molecular analysis of isoleucyl-tRNA synthetase mutations in clinical isolates of methicillin-resistant Staphylococcus aureus with low-level mupirocin resistance.

Yang JA, Park DW, Sohn JW, Yang IS, Kim KH, Kim MJ - J. Korean Med. Sci. (2006)

Bottom Line: All isolates with low-level mupirocin resistance contained the known V588F mutation affecting the Rossman fold, and some of them additionally had previously unidentified mutations such as P187F, K226T, F227L, Q612H, or V767D.Interestingly, Q612H was a novel mutation that was involved in stabilizing the conformation of the catalytic loop containing the KMSKS motif.In conclusion, this study confirms that molecular heterogeneity in ileS gene is common among clinical MRSA isolates with low-level mupirocin resistance, and further study on clinical mutants is needed to understand the structural basis of low-level mupirocin resistance.

View Article: PubMed Central - PubMed

Affiliation: Research Institute of Emerging Infectious Diseases, Korea University, Seoul, Korea.

ABSTRACT
Emergence and spread of low-level mupirocin resistance in staphylococci have been increasingly reported in recent years. The aim of this study was to characterize missense mutations within the chromosomal isoleucyl-tRNA synthetase gene (ileS) among clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) with low-level mupirocin resistance. A total of 20 isolates of MRSA with low-level mupirocin resistance (minimal inhibitory concentration, 16-64 microg/mL) were collected from 79 patients in intensive care units for six months. The isolates were analyzed for isoleucyl-tRNA synthetase (IleS) mutations that might affect the binding of mupirocin to the three-dimensional structure of the S. aureus IleS enzyme. All isolates with low-level mupirocin resistance contained the known V588F mutation affecting the Rossman fold, and some of them additionally had previously unidentified mutations such as P187F, K226T, F227L, Q612H, or V767D. Interestingly, Q612H was a novel mutation that was involved in stabilizing the conformation of the catalytic loop containing the KMSKS motif. In conclusion, this study confirms that molecular heterogeneity in ileS gene is common among clinical MRSA isolates with low-level mupirocin resistance, and further study on clinical mutants is needed to understand the structural basis of low-level mupirocin resistance.

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Structure of IleS from S. aureus Oxford. Distribution of mutated residues identified from MRSA isolates with low-level mupirocin resistance. (A) Overall IleS structure showing bound tRNAIle (violet) and mupirocin (slate/red) (22). The backbone fold of IleS is shown as a ribbon, with individual sites of mutation highlighted displaying 6 residues' side chains (red) and sequence numbers, and yellow-colored structure with the Q612H mutation. (B) Enlarged view of the beta-helix region adjacent to the Rossman fold and residues 588 and 612. Note that the Q612H mutation has potential to interact with mupirocin. Q612 sits close to the core of IleS Rossman fold motif (yellow/blue), where it forms a water-bridged hydrogen bond with the side chain of D635 (arrow).
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Figure 1: Structure of IleS from S. aureus Oxford. Distribution of mutated residues identified from MRSA isolates with low-level mupirocin resistance. (A) Overall IleS structure showing bound tRNAIle (violet) and mupirocin (slate/red) (22). The backbone fold of IleS is shown as a ribbon, with individual sites of mutation highlighted displaying 6 residues' side chains (red) and sequence numbers, and yellow-colored structure with the Q612H mutation. (B) Enlarged view of the beta-helix region adjacent to the Rossman fold and residues 588 and 612. Note that the Q612H mutation has potential to interact with mupirocin. Q612 sits close to the core of IleS Rossman fold motif (yellow/blue), where it forms a water-bridged hydrogen bond with the side chain of D635 (arrow).

Mentions: To better understand the significance of the novel mutations, we used a computer program to superimpose them on the crystal structure of the S. aureus IleS. With the exception of V588F, all the missense mutations we identified in this study are novel mutations. Two of them, V588F and Q612H, are located near the Rossman fold motif, and the others are located outside (Fig. 1A). Because V588 is located in a hydrophobic pocket that interacts directly with the fatty acid side chain of mupirocin, mutation of this residue causes steric repulsion to mupirocin. The Q612H mutation has potential to interact with mupirocin. Q612 sits close to the core of IleS Rossman fold motif, where it forms a water-bridged hydrogen bond with the side chain of D635 (Fig. 1B). Since it is located at about 15 Å from the binding site of mupirocin, it is not clear how the Q612H mutation could directly affect the interaction with mupirocin. The Q612H mutation appeared to affect the catalytic loop configuration during substrate binding, thus leading to the differences in the inhibitory activity of mupirocin.


Molecular analysis of isoleucyl-tRNA synthetase mutations in clinical isolates of methicillin-resistant Staphylococcus aureus with low-level mupirocin resistance.

Yang JA, Park DW, Sohn JW, Yang IS, Kim KH, Kim MJ - J. Korean Med. Sci. (2006)

Structure of IleS from S. aureus Oxford. Distribution of mutated residues identified from MRSA isolates with low-level mupirocin resistance. (A) Overall IleS structure showing bound tRNAIle (violet) and mupirocin (slate/red) (22). The backbone fold of IleS is shown as a ribbon, with individual sites of mutation highlighted displaying 6 residues' side chains (red) and sequence numbers, and yellow-colored structure with the Q612H mutation. (B) Enlarged view of the beta-helix region adjacent to the Rossman fold and residues 588 and 612. Note that the Q612H mutation has potential to interact with mupirocin. Q612 sits close to the core of IleS Rossman fold motif (yellow/blue), where it forms a water-bridged hydrogen bond with the side chain of D635 (arrow).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2721991&req=5

Figure 1: Structure of IleS from S. aureus Oxford. Distribution of mutated residues identified from MRSA isolates with low-level mupirocin resistance. (A) Overall IleS structure showing bound tRNAIle (violet) and mupirocin (slate/red) (22). The backbone fold of IleS is shown as a ribbon, with individual sites of mutation highlighted displaying 6 residues' side chains (red) and sequence numbers, and yellow-colored structure with the Q612H mutation. (B) Enlarged view of the beta-helix region adjacent to the Rossman fold and residues 588 and 612. Note that the Q612H mutation has potential to interact with mupirocin. Q612 sits close to the core of IleS Rossman fold motif (yellow/blue), where it forms a water-bridged hydrogen bond with the side chain of D635 (arrow).
Mentions: To better understand the significance of the novel mutations, we used a computer program to superimpose them on the crystal structure of the S. aureus IleS. With the exception of V588F, all the missense mutations we identified in this study are novel mutations. Two of them, V588F and Q612H, are located near the Rossman fold motif, and the others are located outside (Fig. 1A). Because V588 is located in a hydrophobic pocket that interacts directly with the fatty acid side chain of mupirocin, mutation of this residue causes steric repulsion to mupirocin. The Q612H mutation has potential to interact with mupirocin. Q612 sits close to the core of IleS Rossman fold motif, where it forms a water-bridged hydrogen bond with the side chain of D635 (Fig. 1B). Since it is located at about 15 Å from the binding site of mupirocin, it is not clear how the Q612H mutation could directly affect the interaction with mupirocin. The Q612H mutation appeared to affect the catalytic loop configuration during substrate binding, thus leading to the differences in the inhibitory activity of mupirocin.

Bottom Line: All isolates with low-level mupirocin resistance contained the known V588F mutation affecting the Rossman fold, and some of them additionally had previously unidentified mutations such as P187F, K226T, F227L, Q612H, or V767D.Interestingly, Q612H was a novel mutation that was involved in stabilizing the conformation of the catalytic loop containing the KMSKS motif.In conclusion, this study confirms that molecular heterogeneity in ileS gene is common among clinical MRSA isolates with low-level mupirocin resistance, and further study on clinical mutants is needed to understand the structural basis of low-level mupirocin resistance.

View Article: PubMed Central - PubMed

Affiliation: Research Institute of Emerging Infectious Diseases, Korea University, Seoul, Korea.

ABSTRACT
Emergence and spread of low-level mupirocin resistance in staphylococci have been increasingly reported in recent years. The aim of this study was to characterize missense mutations within the chromosomal isoleucyl-tRNA synthetase gene (ileS) among clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) with low-level mupirocin resistance. A total of 20 isolates of MRSA with low-level mupirocin resistance (minimal inhibitory concentration, 16-64 microg/mL) were collected from 79 patients in intensive care units for six months. The isolates were analyzed for isoleucyl-tRNA synthetase (IleS) mutations that might affect the binding of mupirocin to the three-dimensional structure of the S. aureus IleS enzyme. All isolates with low-level mupirocin resistance contained the known V588F mutation affecting the Rossman fold, and some of them additionally had previously unidentified mutations such as P187F, K226T, F227L, Q612H, or V767D. Interestingly, Q612H was a novel mutation that was involved in stabilizing the conformation of the catalytic loop containing the KMSKS motif. In conclusion, this study confirms that molecular heterogeneity in ileS gene is common among clinical MRSA isolates with low-level mupirocin resistance, and further study on clinical mutants is needed to understand the structural basis of low-level mupirocin resistance.

Show MeSH
Related in: MedlinePlus