Protease-susceptible sites and properties of fragments of aortic smooth-muscle myosin.
We have examined the protease susceptibility of aortic myosin, the thermal unfolding profiles of myosin rod and light meromyosin (LMM) and the solubility properties of the LMM fragments. Two major protease-susceptible sites were found, located at the head-rod junction and the heavy meromyosin (HMM)-LMM junction. Both tryptic and chymotryptic digestion of aortic myosin rod produced the LMM (80-85 kDa) and short subfragment 2 (S-2) (40-45 kDa) segments, which were similar to those of gizzard myosin rod and differed from the short LMM (70 kDa) and long S-2 (58 kDa) segments produced from skeletal-muscle rod. The thermal unfolding profile of aortic myosin rods exhibited three helix-unfolding transitions, at 47.5, 51 and 54 degrees C, similar to those of gizzard rods yet different from those of skeletal-muscle rods. There was a dramatic difference in the solubility of aortic LMM fragments of various molecular mass, as for gizzard smooth-muscle LMM and rabbit skeletal-muscle LMM. LMM fragments of molecular mass 77 kDa or more were completely insoluble in low-ionic-strength buffer, whereas LMM fragments of molecular mass 73 kDa or less were completely soluble in low-ionic-strength buffer. Proteolytic digestion patterns of LMM showed two additional protease-susceptible sites located 13 and 30 kDa from the ends of the LMM molecule. This suggests the existence of flexible regions within the LMM molecule, which may be responsible for the folded form of aortic myosin.
Department of Biochemistry, Chang Gung Medical College, Tao-Yuan, Republic of China.
L
L
King
M J
MJ
Jiang
T S
TS
Huang
G C
GC
Sheu
D000595:Amino Acid Sequence; D000818:Animals; D001011:Aorta / Q000378:metabolism*; D001665:Binding Sites; D002417:Cattle; D002645:Chickens; D002918:Chymotrypsin / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000378:metabolism*; D005895:Gizzard, Avian; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D018482:Muscle, Skeletal / Q000378:metabolism; D009130:Muscle, Smooth / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D015879:Myosin Subfragments / Q000737:chemistry / Q000378:metabolism; D009218:Myosins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D011489:Protein Denaturation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D011817:Rabbits; D013379:Substrate Specificity; D014357:Trypsin / Q000378:metabolism
D015879:Myosin Subfragments; D010446:Peptide Fragments; D010450:Endopeptidases; D002918:Chymotrypsin; D014357:Trypsin; D009218:Myosins