评价农作物蛋白质优劣的一种排序方法──灰色关联度法

依据灰色系统理论中关联度分析法，提出一种对农作物蛋白质氨基酸构成品质进行考察排序的方法，结果与现行的化学评价法具有较好的一致性，且可避免因某一信息的缺乏而使分离结果偏离实际较大的不足。本法也可根据研究目的不同，选取不同的指标总数作为参考数列对待评价样品进行不同角度的评价考察。     

Biochemical identification of residues that discriminate between 3,4-dihydroxyphenylalanine decarboxylase and 3,4-dihydroxyphenylacetaldehyde synthase-mediated reactions

In available insect genomes, there are several L-3,4-dihydroxyphenylalanine (L-dopa) decarboxylase (DDC)-like or aromatic amino acid decarboxylase (AAAD) sequences. This contrasts to those of mammals whose genomes contain only one DDC. Our previous experiments established that two DDC-like proteins from Drosophila actually mediate a complicated decarboxylation-oxidative deamination process of dopa in the presence of oxygen, leading to the formation of 3,4-dihydroxyphenylacetaldehyde (DHPA), CO₂, NH_3, and H₂O₂. This contrasts to the typical DDC-catalyzed reaction, which produces CO₂ and dopamine. These DDC-like proteins were arbitrarily named DHPA synthases based on their critical role in insect soft cuticle formation. Establishment of reactions catalyzed by these AAAD-like proteins solved a puzzle that perplexed researchers for years, but to tell a true DHPA synthase from a DDC in the insect AAAD family remains problematic due to high sequence similarity. In this study, we performed extensive structural and biochemical comparisons between DHPA synthase and DDC. These comparisons identified several target residues potentially dictating DDC-catalyzed and DHPA synthase-catalyzed reactions, respectively. Comparison of DHPA synthase homology models with crystal structures of typical DDC proteins, particularly residues in the active sites, provided further insights for the roles these identified target residues play. Subsequent site-directed mutagenesis of the tentative target residues and activity evaluations of their corresponding mutants determined that active site His192 and Asn192 are essential signature residues for DDC- and DHPA synthase-catalyzed reactions, respectively. Oxygen is required in DHPA synthase-mediated process and this oxidizing agent is reduced to H₂O₂ in the process. Biochemical assessment established that H₂O₂, formed in DHPA synthase-mediated process, can be reused as oxidizing agent and this active oxygen species is reduced to H₂O; thereby avoiding oxidative stress by H₂O₂. Results of our structural and functional analyses provide a reasonable explanation of mechanisms involved in DHPA synthase-mediated reactions. Based on the key active site residue Asn192, identified in Drosophila DHPA synthase, we were able to distinguish all available insect DHPA synthases from DDC sequences primarily.     

Lack of effect of corticotropin releasing factor on hypothalamic dopamine and serotonin synthesis turnover rates in rats

Catecholamine and serotonin neurons in the hypothalamus regulate the secretion of corticotropin releasing factor (CRF). We considered the possibility that CRF might in turn affect the activity of these aminergic neurons. We examined the effect of intracisternal administration of synthetic CRF on the synthesis turnover rates of dopamine and serotonin in the hypothalamus of adult male rats using two different methods to assess turnover. In one study, we measured the accumulation of L-dihydroxyphenylalanine (L-DOPA) or 5-hydroxytryptophan (5-HTP) in mediobasal hypothalamus after L-aromatic amino acid decarboxylase inhibition with m-hydroxybenzylhydrazine 20 min before sacrifice, and in the second study we measured the accumulation of dopamine, norepinephrine, epinephrine and serotonin after monoamine oxidase inhibition with pargyline 20 min before sacrifice. The commercial CRF which we administered intraarterially increased plasma ACTH and corticosterone concentrations. Intracerebral CRF 5 to 20 micrograms 20 min before sacrifice or 20 micrograms 110 min before sacrifice did not alter the m-hydroxybenzylhydrazine-induced accumulation of L-DOPA or 5-HTP when compared with saline vehicle-injected controls. CRF 20 micrograms did not alter basal concentration or pargyline-induced accumulation of the catecholamines or serotonin in whole hypothalamus when compared with saline vehicle-injected controls. Thus, intracisternal administration of CRF did not alter hypothalamic dopamine or serotonin synthesis rates as assessed by two nonsteady state turnover methods. The data suggest that the release of CRF from neurons in hypothalamus does not alter the activity of catecholamine or serotonin neurons in the hypothalamus of normal adult male rats.     

l-2,4-diaminobutyric acid decarboxylase activity responsible for the formation of 1,3-diaminopropane in Enterobacter aerogenes

High content of 1,3-diaminopropane (DAP), a normally minor derivative of polyamine metabolism, have been observed in cells of Enterobacter aerogenes. Supplementation of the growth medium with L-2,4-diaminobutyric acid (L-DABA) resulted in increased production of DAP, but not if supplemented with spermidine. On the basis of these observations, the biosynthetic route for DAP was evaluated. It has appeared that this bacterium possesses a novel enzyme activity catalysing the formation of DAP from L-DABA. Lack of the activity for oxidative cleavage of spermidine yielding DAP suggests that the enzyme termed DABA decarboxylase is responsible for the formation of DAP in this bacterium. The enzyme was partially purified 360-fold and some properties were examined. The pH optimum for the activity was 7.75-8.0, and the enzyme showed an absolute requirement for pyridoxal 5'-phosphate with the Km value of 41 microM. The Km value for L-DABA was 0.32 mM, and neither L-2,3-diaminopropionic acid, L-ornithine nor L-lysine showed detectable substrate activity towards the partially purified enzyme. Mg2+ and dithiothreitol greatly activated the enzyme.