A chromatin-bound proteolytic activity with unique specificity for histone H2A.

A protease associated with purified calf thymus chromatin has been found to act exclusively upon histone H2A, yielding a single new protein species, cH2A. This fragment migrates faster than H2A in acrylamide gel electrophoresis under denaturing conditions. The cH2A was purified and subjected to amino acid analysis and partial sequencing by the use of carboxypeptidase A. These studies demonstrated that cH2A had been derived from the removal of fifteen amino acids from the carboxy-terminal end of the intact H2A molecule, and that valine114 was its new carboxy-terminal residue. This cleavage does not occur under low ionic strength conditions, where H2A is believed to approximate a random coil; rather, it requires high ionic strength conditions similar to those under which the H2A molecule undergoes radical secondary and tertiary structural changes. This dependence upon ionic strength implies that the proteolytic cleavage is conformation- as well as sequence-specific. The H2A-specific protease is of nuclear origin, since isolation of nuclei by methods designed to maximize or minimize cytoplasmic contamination does not affect the level of proteolytic activity associated with purified chromatin. This nuclear protease appears to be tightly associated with the chromatin in vivo, for 0.6 M NaCl will not free it from isolated chromatin. A concentration of 1.2 M NaCl is required to dissociate the protease as well as its substrate from chromatin. The relationship of this enzyme to previously reported chromatin-bound proteases is discussed.