Fragments of beta-galactosidase from Escherichia coli. Fragmentation, purification, characterization and in vitro complementation.

Thermal fragmentation of the beta-galactosidase was studied in different buffer solutions and at different temperatures. Fragmentation of the subunits in small size polypeptides could be observed directly. The fragmentation proceeded in buffer solution, pH 7.0, at either 75 degrees C or 100 degrees C in the presence of sodium dodecyl sulfate. The elevated temperature appeared to accelerate this process. At 100 degrees C, pH 7.2, the fragmentation proceeded in the absence of sodium dodecyl sulfate, but in the presence of 8 M urea. Molecular weights, determined by sodium dodecyl sulfate disc gel electrophoresis were from 130,000 to about 20,000. Multiple bands were observed. After dissociation was complete at 37 degrees C, the fragments were purified by ion-exchange column chromatography. Of the five fragments thus obtained, four were homogeneous by disc-gel electrophoresis. Molecular weight of the homogeneous fragments were found to be near 25,000. The fifth comprised a mixture of four fragments having molecular weights from 29,000 to 72,000. Two of the fragments were active as the alpha-donor in in vitro complementation with mutant M15, which contains a deletion in the alpha-region of the z gene.