Purification, crystallization and preliminary X-ray diffraction analysis of recombinant human neutrophil-activating peptide 2 (rhNAP-2)

FEBS Lett. 1994 Jun 27;347(2-3):300-3. doi: 10.1016/0014-5793(94)00573-7.

Abstract

The potent activator and chemoattractant for human neutrophils, neutrophil-activating peptide 2 (NAP-2), has been cloned and expressed in Escherichia coli. The protein has been purified to homogeneity (> 98%) by a series of chromatographic techniques, including reversed phase HPLC. The biological activity of recombinant human NAP-2 (rhNAP-2), characterized by the induction of elastase release from human neutrophils, was found to be comparable to natural NAP-2. rhNAP-2 has been crystallized by the hanging drop vapor diffusion method. The crystals belong to space group P222 with unit cell dimensions of a = 30.8 A, b = 39.5 A and c = 95.3 A. A packing density of 3.8 A3/Da with a solvent content of approximately 68% is obtained when one molecule per asymmetric unit is assumed. The crystals were shown to diffract to beyond 2.0 A on a conventional X-ray source. They are stable to X-rays for several days and are thus suitable for high resolution structure determination.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Chromatography, High Pressure Liquid
  • Cloning, Molecular
  • Crystallization
  • Crystallography, X-Ray*
  • Electrophoresis, Polyacrylamide Gel
  • Escherichia coli / genetics
  • Gene Expression
  • Humans
  • Leukocyte Elastase
  • Molecular Sequence Data
  • Neutrophils / drug effects
  • Neutrophils / enzymology
  • Pancreatic Elastase / metabolism
  • Peptides / chemistry
  • Peptides / isolation & purification*
  • Peptides / pharmacology
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / isolation & purification
  • beta-Thromboglobulin

Substances

  • PPBP protein, human
  • Peptides
  • Recombinant Proteins
  • beta-Thromboglobulin
  • connective tissue-activating peptide
  • Pancreatic Elastase
  • Leukocyte Elastase