Elsevier

Gene

Volume 113, Issue 1, 1 April 1992, Pages 75-82
Gene

Cloning and sequence determination of a cDNA encoding Aspergillus nidulans calmodulin-dependent multifunctional protein kinase

https://doi.org/10.1016/0378-1119(92)90671-BGet rights and content

Abstract

A partial cDNA encoding Aspergillus nidulans calmodulin-dependent multifunctional protein kinase (ACMPK) was isolated from a λZAP expression library by immunoselection using monospecific polyclonal antibodies to the enzyme. The sequence of both strands of the cDNA (CMKa) was determined. The deduced amino acid (aa) sequence contained all eleven consensus domains found in serine/threonine protein kinases [Hanks et al., Science 241 (1988) 42–52], as well as a putative calmodulin-binding domain. The cDNA contained an intron, lacked an in-frame start codon, and was not polyadenylated. A full-length copy of CMKa was subsequently isolated from a λgt10 library of A. nidulans cDNA using a restriction fragment of the first clone as a probe. It contained an in-frame start codon, an open reading frame (ORF) of 1242 bp and was polyadenylated. The ORF encoded a protein of 414 aa residues with an Mr of 46895 and an isoelectric point pI = 6.4. These values are in good agreement with that observed forthe native enzyme [Bartelt et al., Proc. Natl. Acad. Sci. USA 85 (1988) 3279–3283]. When aligned to optimize homology, 29% of the predicted aa sequence of ACMPK is identical to that of the α-subunit of rat brain calmodulin-dependent protein kinase II. ACMPK shares 40 and 44% identity in aa sequence with YCMK1 and YCMK2, respectively, two Ca2+/calmodulin-dependent protein kinases recently cloned from Saccharomyces cerevisiae [Pausch et al., EMBO J. 10 (1991) 1511–1522]. Results of Southern analysis of restriction digests of genomic DNA indicate that ACMPK is encoded by a single-copy gene.

References (46)

  • C.M. Schworer et al.

    Ca2+/calmodulin-dependent protein kinase, II. Identification of a regulatory autophosphorylation site adjacent to the inhibitory and calmodulin-binding domains

    J. Biol. Chem.

    (1988)
  • E. Southern

    Detection of specific sequences among DNA fragments separated by gel electrophoresis

    J. Mol. Biol.

    (1975)
  • T. Tobimatsu et al.

    Molecular cloning of the cDNA encoding the third polypeptide (7) of brain calmodulin-dependent protein kinase II

    J. Biol. Chem.

    (1988)
  • R.B. Waring et al.

    Characterization of an inducible expression system in Aspergillus nidulans using alcA and tubulin-coding genes

    Gene

    (1989)
  • C. Baitinger et al.

    Multifunctional Ca2+/calmodulin-dependent protein kinase is necessary for nuclear envelope breakdown

    J. Cell Biol.

    (1990)
  • D.C. Bartelt et al.

    Purification, characterization and substrate specificity of calmodulin-dependent myosin light chain kinase from bovine brain

    Biochem. J.

    (1987)
  • D.C. Bartelt et al.

    Calmodulin-dependent multifunctional protein kinase in Aspergillus nidulans

  • D.C. Bartelt et al.

    Expression of cDNA coding for A. nidulans calmodulin-dependent multifunctional protein kinase

    J. Cell Biol.

    (1989)
  • M.K. Bennett et al.

    Deduced primary structure of the β subunit type II Ca2+/calmodulin-dependent protein kinase determined by molecular cloning

  • W.D. Benton et al.

    Screening lambda gt recombinant clones by hybridization to single plaques in situ

    Science

    (1977)
  • S. Chao et al.

    Enhanced specificity in immunoscreening of expression cDNA clones using radiolabelled antigen overlay

    BioTechniques

    (1989)
  • R.A. Dean et al.

    Regulation of the A. nidulans nidulans pectatelyase gene (pelA)

    Plant Cell

    (1989)
  • B.C. Donly et al.

    Frameshift autoregulation in the gene for Escherichia coli release factor 2: partly functional mutants result in frameshift enhancement

    Nucleic Acid Res.

    (1990)
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