EXTENSIVE PROTEOLYTIC DIGESTION OF THE (CA2-ATPASE FROM SARCOPLASMIC-RETICULUM LEADS TO A HIGHLY HYDROPHOBIC - PROTEINACEOUS RESIDUE WITH AMAINLY ALPHA-HELICAL STRUCTURE(+MG2+))

The purified (Ca2++Mg2+)-ATPase from sarcoplasmic reticulum was subjec ted to extensive proteolysis by using trypsin and proteinase K. This d igestion led to the elimination of a considerable portion of the prote in, so that the lipid to protein weight ratio was increased from 0.44 in the purified ATPase to 1.20 after extensive proteolysis. After the digestion, the, residue was found to be considerably enriched in hydro phobic amino acids. FT-IR spectroscopic studies indicated that the sec ondary structure of the proteolytic residue was enriched in alpha-heli x with 75%, compared with 48% in the intact purified ATPase. FT-IR stu dies using ATR polarization showed that the alpha-helical part of the residue of proteolytic digestion was considerably more polarized than the purified ATPase, indicating that, on average, the alpha-helices of the residual protein should lie with an orientation closer to the nor mal to the plane of the membrane. Thermal denaturation studies showed that the residue of proteolysis was considerably more stable than the intact purified ATPase. This would be compatible with the residue bein g protected from denaturation by its hydrophobic location within the m embrane. This study is experimental evidence of the alpha-helical stru cture of the membrane part Of this protein, as suggested by prediction s made from its known primary structure (Brandl et al., 1986).