title: The archaeon Sulfolobus solfataricus contains a membrane-associated protein kinase activity that preferentially phosphorylates threonine residues in vitro.
abstract: The extreme acidothermophilic archaeon Sulfolobus solfataricus harbors a membrane-associated protein kinase activity. Its solubilization and stabilization required detergents, suggesting that this activity resides within an integral membrane protein. The archaeal protein kinase utilized purine nucleotides as phosphoryl donors in vitro. A noticeable preference for nucleotide triphosphates over nucleotide diphosphates and for adenyl nucleotides over the corresponding guanyl ones was observed. The molecular mass of the solubilized, partially purified enzyme was estimated to be approximately 125 kDa by gel filtration chromatography. Catalytic activity resided in a polypeptide with an apparent molecular mass of approximately 67 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Challenges with several exogenous substrates revealed the protein kinase to be relatively selective. Only casein, histone H4, reduced carboxyamidomethylated and maleylated lysozyme, and a peptide modeled after myosin light chains (KKRAARATSNVFA) were phosphorylated to appreciable levels in vitro. All of the aforementioned substrates were phosphorylated on threonine residues, while histone H4 was phosphorylated on serine as well. Substitution of serine for the phosphoacceptor threonine in the myosin light chain peptide produced a noticeably inferior substrate. The protein kinase underwent autophosphorylation on threonine and was relatively insensitive to a set of known inhibitors of "eukaryotic" protein kinases.
authors.affiliation: Department of Biochemistry, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061, USA.
authors.forename: B H
authors.initials: BH
authors.lastname: Lower
authors.forename: K M
authors.initials: KM
authors.lastname: Bischoff
authors.forename: P J
authors.initials: PJ
authors.lastname: Kennelly
mesh_terms: D002462:Cell Membrane / Q000201:enzymology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008670:Metals / Q000378:metabolism; D009711:Nucleotides / Q000378:metabolism; D010455:Peptides / Q000378:metabolism; D010766:Phosphorylation; D010769:Phosphothreonine / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism; D012995:Solubility; D016944:Sulfolobus / Q000201:enzymology* / Q000254:growth & development
chemical_list: D004791:Enzyme Inhibitors; D008670:Metals; D009711:Nucleotides; D010455:Peptides; D011506:Proteins; D010769:Phosphothreonine; D017346:Protein Serine-Threonine Kinases