Purification and amino acid microsequencing of alkaline phosphodiesterase I from calf kidney.
Alkaline phosphodiesterase I was purified from calf kidney following extraction with 0.5% Triton X-100 and then fractionation with a Rotfor cell and high-pressure liquid chromatography. The isoelectric point of alkaline phosphodiesterase I was 5.9. Both the native and the subunit molecular weight of alkaline phosphodiesterase I was determined to be 125 kDa. The N-terminus of its amino acid sequence appeared to be blocked. The immobilized protein, separated by gel electrophoresis and transblotted to a polyvinylidene difluoride membrane, was digested on the membrane by proteases and separated by reverse-phase high-performance liquid chromatography. Eighty one amino acids were sequenced. One of the internal sequences was identical with that of an active-site-containing fragment from calf intestinal enzyme, as reported by Culp et al. (1985).
Maruyama
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E
Division of Mental Retardation and Birth Defect Research, National Institute of Neuroscience, NCNP, Tokyo, Japan.
Iwamatsu
A
A
Takashima
S
S
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D007668:Kidney / Q000201:enzymology*; D008567:Membranes, Artificial; D008969:Molecular Sequence Data; D008970:Molecular Weight; D043264:Phosphodiesterase I; D010727:Phosphoric Diester Hydrolases / Q000737:chemistry / Q000302:isolation & purification*; D011145:Polyvinyls; D005465:Fluorocarbon Polymers
D008567:Membranes, Artificial; D043264:Phosphodiesterase I; D010727:Phosphoric Diester Hydrolases; D011145:Polyvinyls; C024865:polyvinylidene fluoride; D005465:Fluorocarbon Polymers