Characterization of the structure and catalytic activity of Legionella pneumophila VipF.
The pathogenic bacteria Legionella pneumophila is known to cause Legionnaires' Disease, a severe pneumonia that can be fatal to immunocompromised individuals and the elderly. Shohdy et al. identified the L. pneumophila vacuole sorting inhibitory protein VipF as a putative N-acetyltransferase based on sequence homology. We have characterized the basic structural and functional properties of VipF to confirm this original functional assignment. Sequence conservation analysis indicates two putative CoA-binding regions within VipF. Homology modeling and small angle X-ray scattering suggest a monomeric, dual-domain structure joined by a flexible linker. Each domain contains the characteristic beta-splay motif found in many acetyltransferases, suggesting that VipF may contain two active sites. Docking experiments suggest reasonable acetyl-CoA binding locations within each beta-splay motif. Broad substrate screening indicated that VipF is capable of acetylating chloramphenicol and both domains are catalytically active. Given that chloramphenicol is not known to be N-acetylated, this is a surprising finding suggesting that VipF is capable of O-acetyltransferase activity. Proteins 2016; 84:1422-1430. © 2016 Wiley Periodicals, Inc.
Department of Chemistry and Biochemistry, James Madison University, Harrisonburg, Virginia, 22807.
Byron H
BH
Young
Department of Chemistry and Biochemistry, James Madison University, Harrisonburg, Virginia, 22807.
Tracy A
TA
Caldwell
Department of Chemistry and Biochemistry, James Madison University, Harrisonburg, Virginia, 22807.
Aidan M
AM
McKenzie
Department of Chemistry and Biochemistry, James Madison University, Harrisonburg, Virginia, 22807.
Oleksandr
O
Kokhan
Department of Chemistry and Biochemistry, James Madison University, Harrisonburg, Virginia, 22807. berndsce@jmu.edu.
Christopher E
0000-0002-4285-4341
CE
Berndsen
D000105:Acetyl Coenzyme A / Q000737:chemistry* / Q000378:metabolism; D000123:Acetyltransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020134:Catalytic Domain; D002701:Chloramphenicol / Q000737:chemistry* / Q000378:metabolism; D003001:Cloning, Molecular; D017124:Conserved Sequence; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D007700:Kinetics; D016952:Legionella pneumophila / Q000737:chemistry / Q000201:enzymology*; D056004:Molecular Dynamics Simulation; D010957:Plasmids / Q000737:chemistry / Q000378:metabolism; D000072417:Protein Domains; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D040681:Structural Homology, Protein; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
D001426:Bacterial Proteins; D011994:Recombinant Proteins; D002701:Chloramphenicol; D000105:Acetyl Coenzyme A; D000123:Acetyltransferases
acetyltransferase; chloramphenicol; legionella; molecular modeling; small angle X-ray scattering