Proteolysis of paramyosin from Mercenaria mercenaria and properties of its most stable segment.
The helical muscle protein paramyosin appears to consist of three segments of approximately equal size that differ in stability to guanidine hydrochloride and heat. The N-terminal segment is most stable and the C-terminal segment is least stable. These differences in stability serve as a basis for design of proteolytic digestions to specifically remove segments of low and intermediate stability. Thus, at room temperature only the C-terminal region was susceptible to digestion by pepsin or trypsin. Proteolytic removal of the latter region resulted in the accumulation of the remaining two-thirds of the paramyosin molecule as a segment (PPC-1) of 140,000 daltons that was still in a stable helical conformation. Proceeding to more rigorous conditions, papain digestion of either paramyosin or PPC-1 in 4 M guanidine-HCl that would be expected to destabilize all but the N-terminal segment did result in cleavage of all except that region. The N-terminal region accumulated as a helical segment of 74,000 daltons (PPC-2) if digestion was limited to 1.5 hr or a smaller segment of 58,000 daltons (PPC-3) if digestion continued for 24 hr. Stability of the three PPC segments to guanidine-HCl and heat was measured by change in fluorescence of tyrosyl residues upon loss of the helical conformation. The stability of the segments corresponded well with the stability of those regions in the paramyosin molecule from which the segments were believed to have come. Amino acid composition of the PPC segments and of paramyosin were all very similar, and prediction of relative stability of these helical proteins from inspection of gross amino acid composition does not appear promising.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D049872:Bivalvia; D003545:Cysteine / Q000032:analysis; D010450:Endopeptidases; D006146:Guanidines; D006358:Hot Temperature; D008970:Molecular Weight; D009124:Muscle Proteins; D010446:Peptide Fragments / Q000032:analysis*; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence
D000596:Amino Acids; D006146:Guanidines; D009124:Muscle Proteins; D010446:Peptide Fragments; D010450:Endopeptidases; D003545:Cysteine