Fragmin, a microfilament regulatory protein from Physarum polycephalum, is phosphorylated by casein kinase II-type enzymes.
Fragmin is a 42 kDa regulatory protein involved in actin microfilament organization in Physarum polycephalum. We show that fragmin is a target of casein kinase II (CK II) enzymes isolated from evolutionarily divergent species. In Physarum microplasmodia, two such kinases were identified. A serine residue located in the sequence Gly-Gly-Ser-Asp-Leu-Glu constitutes the phosphorylation site and was identified by phosphopeptide sequencing, mass spectometry analysis, and inhibition studies with a synthetic peptide corresponding to this site. Interestingly, the actin-fragmin dimer (A--F) as well as the actin2-fragmin trimer (A2--F) are equally efficient targets, and phosphorylation had no effect on the actin-binding properties of fragmin. Actin-fragmin isolated from microplasmodia revealed a minor acidic fragmin isoform, suggesting that fragmin is phosphorylated in vivo. The actin-fragmin complex is also phosphorylated on the actin subunit by an endogenous actin-fragmin kinase [Gettemans, J., De Ville, Y., Vandekerckhove, J., & Waelkens, E. (1992) EMBO J. 11, 3185-3191]. We show that the two phosphorylation events act independently of each other.
De Corte
V
V
Flanders Interuniversity Institute for Biotechnology, Laboratory for Physiological Chemistry, University of Ghent, Belgium.
Gettemans
J
J
De Ville
Y
Y
Waelkens
E
E
Vandekerckhove
J
J
D000199:Actins / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D047390:Casein Kinase II; D005656:Fungal Proteins / Q000378:metabolism; D008840:Microfilament Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D005810:Multigene Family; D010455:Peptides / Q000737:chemistry; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D010804:Physarum / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*
D000199:Actins; D002352:Carrier Proteins; C034037:F-actin-binding proteins; D005656:Fungal Proteins; D008840:Microfilament Proteins; D010455:Peptides; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates; D010750:Phosphoproteins; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases