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Title: Diacyltransferase Activity and Chain Length Specificity of Mycobacterium tuberculosis PapA5 in the Synthesis of Alkyl β-Diol Lipids

Abstract

Although classified as Gram-positive bacteria, Corynebacterineae possess an asymmetric outer membrane that imparts structural and thereby physiological similarity to more distantly related Gram-negative bacteria. Like lipopolysaccharide in Gram-negative bacteria, lipids in the outer membrane of Corynebacterineae have been associated with the virulence of pathogenic species such as Mycobacterium tuberculosis (Mtb). For example, Mtb strains that lack long, branched-chain alkyl esters known as dimycocerosates (DIMs) are significantly attenuated in model infections. The resultant interest in the biosynthetic pathway of these unusual virulence factors has led to the elucidation of many of the steps leading to the final esterification of the alkyl beta-diol, phthiocerol, with branched-chain fatty acids know as mycocerosates. PapA5 is an acyltransferase implicated in these final reactions. We here show that PapA5 is indeed the terminal enzyme in DIM biosynthesis by demonstrating its dual esterification activity and chain-length preference using synthetic alkyl beta-diol substrate analogues. Applying these analogues to a series of PapA5 mutants, we also revise a model for the substrate binding within PapA5. Finally, we demonstrate that the Mtb Ser/Thr kinase PknB modifies PapA5 on three Thr residues, including two (T196, T198) located on an unresolved loop. These results clarify the DIM biosynthetic pathway and suggest possiblemore » mechanisms by which DIM biosynthesis may be regulated by the post-translational modification of PapA5.« less

Authors:
; ; ; ; ; ; ; ; ; ; ; ;
Publication Date:
Research Org.:
Pacific Northwest National Laboratory (PNNL), Richland, WA (US), Environmental Molecular Sciences Laboratory (EMSL)
Sponsoring Org.:
USDOE
OSTI Identifier:
1243277
Report Number(s):
PNNL-SA-108635
Journal ID: ISSN 0006-2960; 47783; 48680; KP1704020
DOE Contract Number:  
AC05-76RL01830
Resource Type:
Journal Article
Resource Relation:
Journal Name: Biochemistry; Journal Volume: 54; Journal Issue: 35
Country of Publication:
United States
Language:
English
Subject:
Environmental Molecular Sciences Laboratory

Citation Formats

Touchette, Megan H., Bommineni, Gopal R., Delle Bovi, Richard J., Gadbery, John, Nicora, Carrie D., Shukla, Anil K., Kyle, Jennifer E., Metz, Thomas O., Martin, Dwight W., Sampson, Nicole S., Miller, W. T., Tonge, Peter J., and Seeliger, Jessica C.. Diacyltransferase Activity and Chain Length Specificity of Mycobacterium tuberculosis PapA5 in the Synthesis of Alkyl β-Diol Lipids. United States: N. p., 2015. Web. doi:10.1021/acs.biochem.5b00455.
Touchette, Megan H., Bommineni, Gopal R., Delle Bovi, Richard J., Gadbery, John, Nicora, Carrie D., Shukla, Anil K., Kyle, Jennifer E., Metz, Thomas O., Martin, Dwight W., Sampson, Nicole S., Miller, W. T., Tonge, Peter J., & Seeliger, Jessica C.. Diacyltransferase Activity and Chain Length Specificity of Mycobacterium tuberculosis PapA5 in the Synthesis of Alkyl β-Diol Lipids. United States. doi:10.1021/acs.biochem.5b00455.
Touchette, Megan H., Bommineni, Gopal R., Delle Bovi, Richard J., Gadbery, John, Nicora, Carrie D., Shukla, Anil K., Kyle, Jennifer E., Metz, Thomas O., Martin, Dwight W., Sampson, Nicole S., Miller, W. T., Tonge, Peter J., and Seeliger, Jessica C.. Tue . "Diacyltransferase Activity and Chain Length Specificity of Mycobacterium tuberculosis PapA5 in the Synthesis of Alkyl β-Diol Lipids". United States. doi:10.1021/acs.biochem.5b00455.
@article{osti_1243277,
title = {Diacyltransferase Activity and Chain Length Specificity of Mycobacterium tuberculosis PapA5 in the Synthesis of Alkyl β-Diol Lipids},
author = {Touchette, Megan H. and Bommineni, Gopal R. and Delle Bovi, Richard J. and Gadbery, John and Nicora, Carrie D. and Shukla, Anil K. and Kyle, Jennifer E. and Metz, Thomas O. and Martin, Dwight W. and Sampson, Nicole S. and Miller, W. T. and Tonge, Peter J. and Seeliger, Jessica C.},
abstractNote = {Although classified as Gram-positive bacteria, Corynebacterineae possess an asymmetric outer membrane that imparts structural and thereby physiological similarity to more distantly related Gram-negative bacteria. Like lipopolysaccharide in Gram-negative bacteria, lipids in the outer membrane of Corynebacterineae have been associated with the virulence of pathogenic species such as Mycobacterium tuberculosis (Mtb). For example, Mtb strains that lack long, branched-chain alkyl esters known as dimycocerosates (DIMs) are significantly attenuated in model infections. The resultant interest in the biosynthetic pathway of these unusual virulence factors has led to the elucidation of many of the steps leading to the final esterification of the alkyl beta-diol, phthiocerol, with branched-chain fatty acids know as mycocerosates. PapA5 is an acyltransferase implicated in these final reactions. We here show that PapA5 is indeed the terminal enzyme in DIM biosynthesis by demonstrating its dual esterification activity and chain-length preference using synthetic alkyl beta-diol substrate analogues. Applying these analogues to a series of PapA5 mutants, we also revise a model for the substrate binding within PapA5. Finally, we demonstrate that the Mtb Ser/Thr kinase PknB modifies PapA5 on three Thr residues, including two (T196, T198) located on an unresolved loop. These results clarify the DIM biosynthetic pathway and suggest possible mechanisms by which DIM biosynthesis may be regulated by the post-translational modification of PapA5.},
doi = {10.1021/acs.biochem.5b00455},
journal = {Biochemistry},
number = 35,
volume = 54,
place = {United States},
year = {Tue Sep 08 00:00:00 EDT 2015},
month = {Tue Sep 08 00:00:00 EDT 2015}
}