Inhibition of a cold‐active alkaline phosphatase by imipenem revealed by <i>in silico</i> modeling of metallo‐β‐lactamase active sites

Chakraborty, Sandeep; Ásgeirsson, Bjarni; Minda, Renu; Salaye, Lipika; Frère, Jean‐Marie; Rao, Basuthkar J.

doi:10.1016/j.febslet.2012.08.030

Cited by 27 publications

(16 citation statements)

References 53 publications

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“…Previously, we detected a promiscuous serine protease scaffold in APs using CLASP [5], and a scaffold recognizing a β-lactam (imipenem) in a cold-active Vibrio AP [18], [19]. The theoretical foundation of CLASP is that the electrostatic potential difference (EPD) in cognate pairs of active site residues is conserved in proteins with the same functionality.…”

Section: Discussionmentioning

confidence: 99%

“…A recent method employs learning techniques to predict whether proteins have proteolytic activities, but has not identified any novel proteases undetected by other methods [14], [15]. CLASP unraveled a promiscuous serine protease scaffold in alkaline phosphatases (AP) [5], one of the widely studied promiscuous enzyme families [16], [17], and also a scaffold recognizing a β-lactam (imipenem) in a cold-active Vibrio AP [18], [19].…”

Section: Introductionmentioning

confidence: 99%

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A Computational Module Assembled from Different Protease Family Motifs Identifies PI PLC from Bacillus cereus as a Putative Prolyl Peptidase with a Serine Protease Scaffold

et al. 2013

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Proteolytic enzymes have evolved several mechanisms to cleave peptide bonds. These distinct types have been systematically categorized in the MEROPS database. While a BLAST search on these proteases identifies homologous proteins, sequence alignment methods often fail to identify relationships arising from convergent evolution, exon shuffling, and modular reuse of catalytic units. We have previously established a computational method to detect functions in proteins based on the spatial and electrostatic properties of the catalytic residues (CLASP). CLASP identified a promiscuous serine protease scaffold in alkaline phosphatases (AP) and a scaffold recognizing a β-lactam (imipenem) in a cold-active Vibrio AP. Subsequently, we defined a methodology to quantify promiscuous activities in a wide range of proteins. Here, we assemble a module which encapsulates the multifarious motifs used by protease families listed in the MEROPS database. Since APs and proteases are an integral component of outer membrane vesicles (OMV), we sought to query other OMV proteins, like phospholipase C (PLC), using this search module. Our analysis indicated that phosphoinositide-specific PLC from Bacillus cereus is a serine protease. This was validated by protease assays, mass spectrometry and by inhibition of the native phospholipase activity of PI-PLC by the well-known serine protease inhibitor AEBSF (IC50 = 0.018 mM). Edman degradation analysis linked the specificity of the protease activity to a proline in the amino terminal, suggesting that the PI-PLC is a prolyl peptidase. Thus, we propose a computational method of extending protein families based on the spatial and electrostatic congruence of active site residues.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

A Computational Module Assembled from Different Protease Family Motifs Identifies PI PLC from Bacillus cereus as a Putative Prolyl Peptidase with a Serine Protease Scaffold

et al. 2013

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“…We have in the past used electrostatic congruence to detect a promiscuous serine protease scaffold in alkaline phosphatases 40 and a phosphoinositide-specific phospholipase C from Bacillus cereus 41 , and a scaffold recognizing a β -lactam (imipenem) in a cold-active Vibrio alkaline phosphatase 42, 43 . However, continuum models 44 that compute potential differences and pK a values from charge interactions in proteins 45 are sensitive to the spatial arrangement of the atoms in the structure.…”

Section: Resultsmentioning

confidence: 99%

Protein structure quality assessment based on the distance profiles of consecutive backbone Cα atoms

et al. 2013

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“…Previous work by our group has established the spatial and electrostatic congruence in cognate residue pairs of the active site in proteins with the same functionality (CLASP) 20, 21 . CLASP analysis indicated that the phosphoinositide-specific phospholipase C (PI-PLC) from Bacillus cereus has spatial and electrostatic congruence with a serine protease motif 22 .…”

Section: Introductionmentioning

confidence: 99%

Dipeptidyl peptidase-IV inhibitors used in type-2 diabetes inhibit a phospholipase C: a case of promiscuous scaffolds in proteins

Chakraborty¹,

Rendón-Ramírez²,

Ásgeirsson³

et al. 2015

F1000Res

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The long term side effects of any newly introduced drug is a subject of intense research, and often raging controversies. One such example is the dipeptidyl peptidase-IV (DPP4) inhibitor used for treating type 2 diabetes, which is inconclusively implicated in increased susceptibility to acute pancreatitis. Previously, based on a computational analysis of the spatial and electrostatic properties of active site residues, we have demonstrated that phosphoinositide-specific phospholipase C (PI-PLC) from Bacillus cereus is a prolyl peptidase using in vivo experiments. In the current work, we first report the inhibition of the native activity of PI-PLC by two DPP4 inhibitors - vildagliptin (LAF-237) and K-579. While vildagliptin inhibited PI-PLC at micromolar concentrations, K-579 was a potent inhibitor even at nanomolar concentrations. Subsequently, we queried a comprehensive, non-redundant set of 5000 human proteins (50% similarity cutoff) with known structures using serine protease (SPASE) motifs derived from trypsin and DPP4. A pancreatic lipase and a gastric lipase are among the proteins that are identified as proteins having promiscuous SPASE scaffolds that could interact with DPP4 inhibitors. The presence of such scaffolds in human lipases is expected since they share the same catalytic mechanism with PI-PLC. However our methodology also detects other proteins, often with a completely different enzymatic mechanism, that have significantly congruent domains with the SPASE motifs. The reported elevated levels of serum lipase, although contested, could be rationalized by inhibition of lipases reported here. In an effort to further our understanding of the spatial and electrostatic basis of DPP4 inhibitors, we have also done a comprehensive analysis of all 76 known DPP4 structures liganded to inhibitors till date. Also, the methodology presented here can be easily adopted for other drugs, and provide the first line of filtering in the identification of pathways that might be inadvertently affected due to promiscuous scaffolds in proteins.

show abstract

Inhibition of a cold‐active alkaline phosphatase by imipenem revealed by in silico modeling of metallo‐β‐lactamase active sites

Cited by 27 publications

References 53 publications

A Computational Module Assembled from Different Protease Family Motifs Identifies PI PLC from Bacillus cereus as a Putative Prolyl Peptidase with a Serine Protease Scaffold

A Computational Module Assembled from Different Protease Family Motifs Identifies PI PLC from Bacillus cereus as a Putative Prolyl Peptidase with a Serine Protease Scaffold

Protein structure quality assessment based on the distance profiles of consecutive backbone Cα atoms

Dipeptidyl peptidase-IV inhibitors used in type-2 diabetes inhibit a phospholipase C: a case of promiscuous scaffolds in proteins

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