Crystal structure and tartrate inhibition of Legionella pneumophila histidine acid phosphatase

• Published in: Archives of biochemistry and biophysics 2015-11, Vol.585, p.32-38
• Main Authors: Dhatwalia, Richa, Singh, Harkewal, Reilly, Thomas J., Tanner, John J.
• Format: Article
• Language: English
• Subjects: Acid Phosphatase - antagonists & inhibitors; Acid Phosphatase - chemistry; Acid Phosphatase - genetics; Adenosine Monophosphate - chemistry; Amino Acid Motifs; Bacterial Proteins - antagonists & inhibitors; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Catalytic Domain; Crystallography, X-Ray; Escherichia coli - genetics; Escherichia coli - metabolism; Gene Expression; Histidine - chemistry; Histidine acid phosphatase; Histidine phosphatase superfamily; Kinetics; Legionella pneumophila - chemistry; Legionella pneumophila - enzymology; Models, Molecular; Molecular Sequence Data; Protein Structure, Secondary; Protein Structure, Tertiary; Recombinant Proteins - chemistry; Recombinant Proteins - genetics; Static Electricity; Substrate recognition; Substrate Specificity; Tartrates - chemistry; X-ray crystallography
• ISSN: 0003-9861; 1096-0384
• DOI: 10.1016/j.abb.2015.09.010

Histidine acid phosphatases (HAPs) utilize a nucleophilic histidine residue to catalyze the transfer of a phosphoryl group from phosphomonoesters to water. HAPs function as protein phosphatases and pain suppressors in mammals, are essential for Giardia lamblia excystation, and contribute to virulence of the category A pathogen Francisella tularensis. Herein we report the first crystal structure and steady-state kinetics measurements of the HAP from Legionella pneumophila (LpHAP), also known as Legionella major acid phosphatase. The structure of LpHAP complexed with the inhibitor l(+)-tartrate was determined at 2.0 Å resolution. Kinetics assays show that l(+)-tartrate is a 50-fold more potent inhibitor of LpHAP than of other HAPs. Electrostatic potential calculations provide insight into the basis for the enhanced tartrate potency: the tartrate pocket of LpHAP is more positive than other HAPs because of the absence of an ion pair partner for the second Arg of the conserved RHGXRXP HAP signature sequence. The structure also reveals that LpHAP has an atypically expansive active site entrance and lacks the nucleotide substrate base clamp found in other HAPs. These features imply that nucleoside monophosphates may not be preferred substrates. Kinetics measurements confirm that AMP is a relatively inefficient in vitro substrate of LpHAP. [Display omitted] •First structure of Legionella major acid phosphatase, a histidine acid phosphatase.•Unpaired Arg accounts for high potency of the inhibitor l(+)-tartrate.•Atypical features include expansive active site and absence of a nucleotide clamp.•Structure suggests that nucleoside monophosphates are not preferred substrates.•Kinetics measurements confirm that AMP is a relatively poor substrate.