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. 2013 Apr;68(4):800-5.
doi: 10.1093/jac/dks497. Epub 2012 Dec 11.

Unexpected N-acetylation of capreomycin by mycobacterial Eis enzymes

Jacob L Houghton  1 , Keith D Green, Rachel E Pricer, Abdelrahman S Mayhoub, Sylvie Garneau-Tsodikova
Affiliations

Unexpected N-acetylation of capreomycin by mycobacterial Eis enzymes

Jacob L Houghton et al. J Antimicrob Chemother. 2013 Apr.

Abstract

Objectives: The enhanced intracellular survival (Eis) protein from Mycobacterium tuberculosis (Eis_Mtb), a regio-versatile N-acetyltransferase active towards many aminoglycosides (AGs), confers resistance to kanamycin A in some cases of extensively drug-resistant tuberculosis (XDR-TB). We assessed the activity of Eis_Mtb and of its homologue from Mycobacterium smegmatis (Eis_Msm) against a panel of anti-tuberculosis (TB) drugs and lysine-containing compounds.

Methods and results: Both enzymes acetylated capreomycin and some lysine-containing compounds, but not other non-AG non-lysine-containing drugs tested. Modelling studies predicted the site of modification on capreomycin to be one of the two primary amines in its β-lysine side chain. Using Eis_Mtb, we established via nuclear magnetic resonance (NMR) spectroscopy that acetylation of capreomycin occurs on the ε-amine of the β-lysine side chain. Using Msm, we also demonstrated for the first time to our knowledge that acetylation of capreomycin results in deactivation of the drug.

Conclusions: Eis is a unique acetyltransferase capable of inactivating the anti-TB drug capreomycin, AGs and other lysine-containing compounds.

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Figures

Figure 1.
Figure 1.
6′-N-acetylation of capreomycin (CAP) by Eis. (a) Scheme showing the conversion of CAP IA and CAP IB into 6′-N-acetyl-CAP IA (Ac-CAP IA) and 6′-N-acetyl-CAP IB (Ac-CAP IB), respectively, by Eis_Mtb. (b) TLC showing the Ac-CAP IA and Ac-CAP IB products (left-hand lane) generated by Eis_Mtb using CAP (mixture of CAP IA and CAP IB) (right-hand lane) with 2.5 eq of AcCoA. (c) Spectrophotometric assay plot monitoring the conversion of CAP into Ac-CAP by Eis_Mtb (black circles) and Eis_Msm (grey circles). (d) Mass spectrum of Ac-CAP (a mixture of Ac-CAP IA and Ac-CAP IB). (e) Michaelis–Menten analysis of the Eis_Mtb-catalysed N-acetylation of CAP.
Figure 2.
Figure 2.
Spectrophotometric assay plots monitoring the acetylation of (a) lysine (circles), di-lysine (triangles), tri-lysine (squares) and tetra-lysine (diamonds) by Eis_Mtb, as well as of (b) neurogranin (triangles). (c) Michaelis–Menten analysis of the Eis_Mtb catalysed N-acetylation of tetra-lysine.
See this image and copyright information in PMC

References

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