Purification, characterization and structure of protein phosphatase 1 from the cilia of Paramecium tetraurelia.
A type 1 serine/threonine protein phosphatase (PP1) which is mostly localized in the excitable ciliary membranes from the protozoan Paramecium, was purified to homogeneity. Approximately 4 micrograms enzyme of 37 kDa was isolated from 100 l axenic culture. The enzymic properties were characterized using phosphorylase a from rabbit skeletal muscle as a substrate and several known effectors of mammalian PP1. The protozoan PP1 was enzymically indistinguishable from its mammalian congener. The amino acid sequence of the Paramecium PP1 was deduced from its cDNA. The full-length clone was obtained in several steps starting with a pair of degenerate primers made according to the two most conserved peptides of rabbit PP1 and PP2A. The gene encodes a protein of 36,392 Da. The identity of the cloned gene and the isolated ciliary PP1 was unequivocally established by microsequencing of four tryptic and cyanogen-bromide peptides which were generated from the purified protein. Paramecium PP1 shows 75% amino-acid-sequence identity with rabbit PP1 alpha. Areas of major differences are the C-termini and N-termini and a sequence between residues 219-242.
Friderich
G
G
Pharmazeutisches Institut, Universität Tübingen, Federal Republic of Germany.
Klumpp
S
S
Russell
C B
CB
Hinrichsen
R D
RD
Kellner
R
R
Schultz
J E
JE
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002923:Cilia / Q000201:enzymology*; D003001:Cloning, Molecular; D004247:DNA / Q000737:chemistry; D008969:Molecular Sequence Data; D009132:Muscles / Q000201:enzymology; D010247:Paramecium / Q000201:enzymology*; D010749:Phosphoprotein Phosphatases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D010762:Phosphorylase a / Q000378:metabolism; D054645:Protein Phosphatase 1; D011817:Rabbits; D013379:Substrate Specificity
D004247:DNA; D010762:Phosphorylase a; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1