Receptor-like protein tyrosine phosphatases generally contain one or two conserved intracellular catalytic domains with a conserved sequence motif ([I/V]HCXAGXXR[S/T]G), a single transmembrane domain, and an external highly variable part. Here, we describe cloning of the intracellular catalytic domain of the rat protein tyrosine phosphatase eta (rPTP eta CD) into pET28a(+) vector, its expression in Escherichia coli, purification and initial characterization. The purification of His(6)-tagged rPTP eta CD to near homogeneity was achieved by a combination of affinity and size exclusion chromatography. The His-tag was subsequently removed by thrombin digestion. PhastGel IEF electrophoresis demonstrated that the isoelectric point of this 41 kDa His(6)-tag free recombinant protein was 7.3, which is just slightly higher than the theoretically predicted value of 7.2. To assess the functionality of the rPTP eta CD we used the pNPP hydrolysis assay and observed that the enzyme has a specific activity of 9 nmol/min/mu g. The secondary structure and stability of the recombinant protein was also analyzed by circular dichroism and fluorescence spectroscopy. In summary, the rPTP eta CD is stable at 18 degrees C, properly folded, and fully active, which makes it a suitable candidate for structural and functional studies. (c) 2005 Elsevier Inc. All rights reserved.