The protein-tyrosine kinases (PTKs) are a burgeoning family of proteins, each of which bears a conserved domain of 250 to 300 amino acids capable of phosphorylating substrate proteins on tyrosine residues. We recently exploited the existence of two highly conserved sequence elements within the catalytic domain to generate PTK-specific degenerate oligonucleotide primers (A. F. Wilks, Proc. Natl. Acad. Sci. USA 86: [1603][1604][1605][1606][1607] 1989). By application of the polymerase chain reaction, portions of the catalytic domains of several novel PTKs were amplified. We describe here the primary sequence of one of these new PTKs, JAK1 (from Janus kinase), a member of a new class of PTK characterized by the presence of a second phosphotransferase-related domain immediately N terminal to the PTK domain. The second phosphotransferase domain bears all the hallmarks of a protein kinase, although its structure differs significantly from that of the PTK and threonine/ serine kinase family members. A second member of this family (JAK2) has been partially characterized and exhibits a similar array of kinase-related domains. JAK1 is a large, widely expressed membrane-associated phosphoprotein of approximately 130,000 Da. The PTK activity of JAK1 has been located in the C-terminal PTK-like domain. The role of the second kinaselike domain is unknown.Protein-tyrosine kinases (PTKs) are structurally well suited to a role in intracellular signal transduction. Many growth factor receptors, for example, transduce the extracellular stimulus they receive through interaction with their cognate ligand via an intracellular tyrosine kinase domain (5, 33, 52; reviewed in reference 60). Members of the PTK family each bear a highly related "catalytic" domain. The phylogenetic relationships established by an amino acid sequence comparison of the catalytic domains (10) are borne out in the overall structure of the PTKs. For example, families of PTKs, such as those based on the structure of the colony-stimulating factor-1 growth factor receptor (38) (including the two types of the platelet-derived growth factor receptor [4,58]) and the protooncogene c-kit [59]) and those clustered around the cytoplasmic PTKs c-src (29) (including HCK/bmk [12], LCK [28], and c-yes [42], among others) and c-fes (37) (including c-FER/flk [11,25]) each share the highest degree of identity with other members of their cluster and, in respect to their overall topology, are structurally more related to each other than to members of other classes of PTK. Hence, the recombination of the PTK catalytic domain with a wide variety of regulatory and other interactive domains suggests a strong evolutionary drive toward the rapid expansion of the use of its physiologically powerful catalytic activity. This combinatorial approach to the evolution of multidomain proteins such as the PTIK family predicts the extensive utilization of the basic tyrosine kinase domain in other metabolic niches.Application of the polymerase chain reaction (PCR) (32, 40) using degenerate PTK-spec...
The novel COMT inhibitor tolcapone was well tolerated at oral doses of 5 to 800 mg. Tolcapone concentration-dependently inhibited COMT activity in erythrocytes and exhibited dose-proportional kinetics. Further investigations into its applicability in the treatment of Parkinson's disease are warranted.
1 Single oral doses of the catechol-O-methyltransferase (COMT) inhibitor tolcapone (10-800 mg) or placebo were administered simultaneously with a dose of levodopa/benserazide 100/25 mg to seven sequential groups of six healthy male subjects in a two-way crossover study. 2 Plasma concentrations of tolcapone, its metabolite 3-O-methyltolcapone, levodopa and 3-O-methyldopa (3-OMD) were determined in conjunction with COMT activity in erythrocytes. 3 The drug combination was well tolerated at all dose levels and there were no signs indicative of an increase in dopaminergic stimulation. 4 Tolcapone caused a rapid and reversible inhibition of COMT activity in erythrocytes in parallel with a dose-dependent decrease in the formation of 3-OMD. Tolcapone increased the area under the concentration-time curve and elimination half-life of levodopa. The maximum effects were obtained at a dose of about 200 mg when both parameters increased approximately twofold. The drug had no influence on the maximum concentration of levodopa. 5 Tolcapone was rapidly absorbed and eliminated with, on average, a tmax of 1.5 h and a ti12 of 2.3 h. The drug showed dose-proportional pharmacokinetics, in contrast to 3-O-methyltolcapone whose formation was relatively decreased at higher doses. 6 Plasma concentrations of tolcapone correlated with inhibition of COMT activity in erythrocytes and suppression of 3-OMD levels, but not with changes in levodopa pharmacokinetics.
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