Preliminary crystallographic characterization of BSAP, an extracellular aminopeptidase fromBacillus subtilis
| Autor: | Yuval Shoham, Gil Shoham, R. Gilboa, Yifat Fundoiano-Hershcovitz, Gali Golan, V. Reiland |
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| Rok vydání: | 2004 |
| Předmět: |
Peptide
Bacillus subtilis Biology Crystallography X-Ray medicine.disease_cause Aminopeptidases Aminopeptidase Catalysis law.invention chemistry.chemical_compound X-Ray Diffraction Structural Biology law Catalytic Domain Escherichia coli medicine Crystallization chemistry.chemical_classification X-Rays General Medicine Hydrogen-Ion Concentration biology.organism_classification Recombinant Proteins Protein Structure Tertiary Amino acid Zinc Crystallography Monomer Enzyme chemistry Peptides |
| Zdroj: | Acta Crystallographica Section D Biological Crystallography. 60:2371-2376 |
| ISSN: | 0907-4449 |
| DOI: | 10.1107/s0907444904026976 |
| Popis: | The extracellular aminopeptidase from Bacillus subtilis (BSAP) has recently been cloned, overexpressed and purified from Escherichia coli. It is a monomer with a molecular weight of 46 425 Da, consisting of 425 amino-acid residues and a double-zinc catalytic centre. The recombinant enzyme was found to be stable for 20 min at 353 K, to function optimally in the pH range 8-9 and to prefer basic and large hydrophobic N-terminal amino acids in peptide and protein substrates. As such, this enzyme can be used as a representative model for structural, functional and mechanistic studies of monomeric double-zinc aminopeptidases, many of which have been found to be involved in medically important biological activities. In this report, the crystallization and preliminary crystallographic characterization of wild-type BSAP are described. Two different crystal forms are reported, of which the hexagonal form H2 is the more suitable for structural study, with average unit-cell dimensions a = b = 226.5, c = 42.8 A. A full diffraction data set has been collected from such a crystal of the native enzyme (2.2 A resolution, 91.2% completeness, R(merge) = 7.1%). A multiwavelength anomalous diffraction (MAD) data set was collected on native (zinc-containing) BSAP at three wavelengths around the zinc absorption edge (peak data set at 2.5 A resolution, 98.8% completeness, R(merge) = 5.3%). These diffraction data were collected at 95-100 K using a synchrotron X-ray source and a CCD area detector. The data are currently being used to obtain crystallographic phasing and to determine the detailed three-dimensional structure of the enzyme. |
| Databáze: | OpenAIRE |
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