Characterization of rat pulmonary monoamine oxidase.

The substrate specificities and kinetics of rat lung monoamine oxidase (MAO) have been studied. Utilizing the irreversible MAO inhibitors, clorgyline and deprenyl, rat lung was shown to possess at least two types of MAO, A and B. Tyramine was a substrate for both forms of the enzyme, whereas 5-hydroxytryptamine (5-HT) was a preferred substrate for the A-form. In contrast to most other tissues, 2-phenylethylamine was not solely a B-type substrate for the rat lung MAO and some metabolism by the A-type was apparent $\text{(}\text{B}\text{A}\text{=}\text{80}\text{20}\text{)}$. Using tyramine as substrate the ratio A/B was shown to be $\text{55}\text{45}$. Rat pulmonary MAO-B was inhibited by deprenyl and the kinetics of MAO-A studied. The Km values for the A-form for tyramine, phenylethylamine and 5-HT were relatively similar and were 270, 244 and 170 μM respectively. Whereas, when the A-form was inhibited by clorgyline, the Km values for the B-form were found to differ considerably: 330, 42 and 850 μM for tyramine, phenylethylamine and 5-HT respectively.     

Tetrahydroisoquinoline 1-carboxamides as growth hormone secretagogues

Several novel series of tetrahydroisoquinoline 1-carboxamides were prepared and shown to be potent growth hormone (GH) secretagogues. Among them, carbamate 12a-E2 displays excellent in vivo activity by increasing plasma GH 10-fold in an anesthetized IV rat model.     

Proteolytic activation of ETK/Bmx tyrosine kinase by caspases

Etk/Bmx is a member of the Btk/Tec family of kinases, which are characterized by having a pleckstrin homology domain at the N terminus, in addition to the Src homology 3 (SH3), SH2, and the catalytic domains, shared with the Src family kinases. Etk, or Btk kinases in general, has been implicated in the regulation of apoptosis. To test whether Etk is the substrate for caspases during apoptosis, in vitro translated [(35)S]methionine-labeled Etk was incubated with different apoptotic extracts and recombinant caspases, respectively. Results showed that Etk was proteolyzed in all conditions tested with identical cleavage patterns. Caspase-mediated cleavage of Etk generated a C-terminal fragment, containing the complete SH2 and tyrosine kinase domains, but without intact pleckstrin homology and SH3 domains. This fragment has 4-fold higher kinase activity than that of the full-length Etk. Ectopic expression of the C-terminal fragment of Etk sensitized the PC3 prostate cancer cells to apoptosis in response to apoptosis-inducing stimuli. The finding, together with an earlier report that Etk is potentially antiapoptotic, suggests that Etk may serve as an apoptotic switch, depending on the forms of Etk existing inside the cells. To our knowledge, this is the first case where the activity of a tyrosine kinase is induced by caspase cleavage.     

A Note on the Log‐Rank Test in Life Table Analysis with Correlated Observations

Survival data consisting of independent sets of correlated failure times may arise in many situations. For example, we may take repeated observations of the failure time of interest from each patient or observations of the failure time on siblings, or consider the failure times on littermates in toxicological experiments. Because the failure times taken on the same patient or related family members or from the same litter are likely correlated, use of the classical log‐rank test in these situations can be quite misleading with respect to type I error. To avoid this concern, this paper develops two closed‐form asymptotic summary tests, that account for the intraclass correlation between the failure times within patients or units. In fact, one of these two test includes the classical log‐rank test as a special case when the intraclass correlation equals 0. Furthermore, to evaluate the finite‐sample performance of the two tests developed here, this paper applies Monte Carlo simulation and notes that they can actually perform quite well in a variety of situations considered here.     

Identification and characterization of a succinyl-coenzyme A (CoA):benzoate CoA transferase in Geobacter metallireducens

Geobacter metallireducens is a Fe(III)-respiring deltaproteobacterium and serves as a model organism for aromatic compound-degrading, obligately anaerobic bacteria. In this study, a genetic system was established for G. metallireducens using nitrate as an alternative electron acceptor. Surprisingly, disruption of the benzoate-induced bamY gene, encoding a benzoate coenzyme A (CoA) ligase, reproducibly showed an increased biomass yield in comparison to the wild type during growth with benzoate but not during growth with acetate. Complementation of bamY in trans converted the biomass yield back to the wild-type level. Growth of the bamY mutant with benzoate can be rationalized by the identification of a previously unknown succinyl-CoA:benzoate CoA transferase activity; it represents an additional, energetically less demanding mode of benzoate activation. The activity was highly enriched from extracts of cells grown on benzoate, yielding a 50-kDa protein band; mass spectrometric analysis identified the corresponding benzoate-induced gene annotated as a CoA transferase. It was heterologously expressed in Escherichia coli and characterized as a specific succinyl-CoA:benzoate CoA transferase. The newly identified enzyme in conjunction with a benzoate-induced succinyl-CoA synthetase links the tricarboxylic acid cycle to the upper benzoyl-CoA degradation pathway during growth on aromatic growth substrates.