Crystallization and preliminary X‐ray diffraction analysis of proline iminopeptidase from <i>Xanthomonas campestris</i> pv. <i>citri</i>

Medrano, Francisco J.; Alonso, Jorge; Garcı́a, José Luis; Bode, Wolfram; Gomis-Rüth, F.X.

doi:10.1016/s0014-5793(96)01363-4

Cited by 5 publications

(5 citation statements)

References 25 publications

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“…XCPIP was purified and crystallized using NaCl as precipitating agent as previously described (Medrano et al ., 1997). These crystals belong to the orthorhombic space group C222, contain one homodimer per asymmetric unit and have cell constants a = 147.2 Å, b = 167.8 Å and c = 85.6 Å.…”

Section: Methodsmentioning

confidence: 99%

Structure of proline iminopeptidase from Xanthomonas campestris pv. citri: a prototype for the prolyl oligopeptidase family

Medrano¹,

Alonso

Garcı́a

et al. 1998

The EMBO Journal

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The proline iminopeptidase from Xanthomonas campestris pv. citri is a serine peptidase that catalyses the removal of N-terminal proline residues from peptides with high specificity. We have solved its threedimensional structure by multiple isomorphous replacement and refined it to a crystallographic R-factor of 19.2% using X-ray data to 2.7 Å resolution. The protein is folded into two contiguous domains. The larger domain shows the general topology of the α/β hydrolase fold, with a central eight-stranded β-sheet flanked by two helices and the 11 N-terminal residues on one side, and by four helices on the other side. The smaller domain is placed on top of the larger domain and essentially consists of six helices. The active site, located at the end of a deep pocket at the interface between both domains, includes a catalytic triad of Ser110, Asp266 and His294. Cys269, located at the bottom of the active site very close to the catalytic triad, presumably accounts for the inhibition by thiolspecific reagents. The overall topology of this iminopeptidase is very similar to that of yeast serine carboxypeptidase. The striking secondary structure similarity to human lymphocytic prolyl oligopeptidase and dipeptidyl peptidase IV makes this proline iminopeptidase structure a suitable model for the three-dimensional structure of other peptidases of this family.

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Section: Methodsmentioning

confidence: 99%

Structure of proline iminopeptidase from Xanthomonas campestris pv. citri: a prototype for the prolyl oligopeptidase family

Medrano¹,

Alonso

Garcı́a

et al. 1998

The EMBO Journal

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show abstract

“…The first and largest is very similar to the R/β-hydrolase fold found in yeast serine carboxypeptidase, and the second is placed on top of the larger domain and essentially consists of six helices. 17 This enzyme is a model for the Pro oligopeptidase folding family.…”

Section: B Structures and Foldsmentioning

confidence: 99%

Irreversible Inhibitors of Serine, Cysteine, and Threonine Proteases

et al. 2002

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“…Structure determination of GaPIP was performed by molecular replacement using X. campestris PIP structure (PDB ID: 1AZW; [29]) and this structure has been deposited in PDB (PDB ID: 5YHP) (Figure 7). It was also shown that GaPIP has nine clefts and two pores.…”

Section: Structure Determination Of Gapipmentioning

confidence: 99%

Biochemical Characterisation and Structure Determination of a Novel Cold-Active Proline Iminopeptidase from the Psychrophilic Yeast, Glaciozyma antarctica PI12

et al. 2022

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Microbial proteases constitute one of the most important groups of industrially relevant enzymes. Proline iminopeptidases (PIPs) that specifically release amino-terminal proline from peptides are of major interest for applications in food biotechnology. Proline iminopeptidase has been extensively characterised in bacteria and filamentous fungi. However, no similar reports exist for yeasts. In this study, a protease gene from Glaciozyma antarctica designated as GaPIP was cloned and overexpressed in Escherichia coli. Sequence analyses of the gene revealed a 960 bp open reading frame encoding a 319 amino acid protein (35,406 Da). The purified recombinant GaPIP showed a specific activity of 3561 Umg−1 towards L-proline-p-nitroanilide, confirming its identity as a proline iminopeptidase. GaPIP is a cold-active enzyme with an optimum activity of 30 °C at pH 7.0. The enzyme is stable between pH 7.0 and 8.0 and able to retain its activity at 10–30 °C. Although GaPIP is a serine protease, only 25% inhibition by the serine protease inhibitor, phenylmethanesulfonylfluoride (PMSF) was recorded. This enzyme is strongly inhibited by the presence of EDTA, suggesting that it is a metalloenzyme. The dimeric structure of GaPIP was determined at a resolution of 2.4 Å. To date, GaPIP is the first characterised PIP from yeasts and the structure of GaPIP is the first structure for PIP from eukaryotes.

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Crystallization and preliminary X‐ray diffraction analysis of proline iminopeptidase from Xanthomonas campestris pv. citri

Cited by 5 publications

References 25 publications

Structure of proline iminopeptidase from Xanthomonas campestris pv. citri: a prototype for the prolyl oligopeptidase family

Structure of proline iminopeptidase from Xanthomonas campestris pv. citri: a prototype for the prolyl oligopeptidase family

Irreversible Inhibitors of Serine, Cysteine, and Threonine Proteases

Biochemical Characterisation and Structure Determination of a Novel Cold-Active Proline Iminopeptidase from the Psychrophilic Yeast, Glaciozyma antarctica PI12

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