PURIFICATION AND MICROSEQUENCING OF HYALURONIDASE ISOZYMES FROM HUMANURINE

We recently cloned the major hyaluronidase of human plasma, which we t ermed HYAL1, All hyaluronidase activity could be purified from human u rine on an anti-HYAL1 monoclonal antibody column, However, urine conta ins two hyaluronidases of 57 kDa and 45 kDa, whereas plasma only conta ins the 57 kDa activity, Microsequencing confirmed that both urinary i sozymes have N-termini identical to plasma hyaluronidase, but a second N-terminus was found in the smaller isozyme, apparently derived from the terminal 25 amino acids of HYAL1, at the C-terminus, The two polyp eptides of the 45 kDa isozyme resulting from endoproteolytic cleavage of the 57 kDa isoform are presumably linked by disulfide bonds, Sperm contains two isozymes of the testicular hyaluronidase, PH-20, and the lower molecular weight isozyme is believed to be an endoproteolyticall y processed form of the larger protein, Analogously to PH-20, the smal ler isozyme of HYAL1 is likely to be a proteolytically processed produ ct of the larger isozyme. (C) 1997 Federation of European Biochemical Societies.