水稻巯基蛋白酶抑制剂的分子修饰与其对稻瘟病菌的抑制作用

水稻巯基蛋白酶抑制剂（ＣＰＩ）经用二硫苏糖醇,对氯汞苯甲酸和碘乙酸修饰后,对木瓜蛋白酶的抑制活性并无改变;用Ｎ－乙基顺丁烯二酰亚胺与ＣＰＩ反应,可以测出ＣＰＩ分子内有１９个巯基被修饰,被修饰后,抑制活性仍无改变,表明水稻ＣＰＩ的抑制活性不需要巯基参与;应用Ｎ－溴代丁二酰亚胺与ＣＰＩ反应,可测出ＣＰＩ分子内有２个Ｔｒｐ被修饰,修饰后,抑制活性全部丧失,表明Ｔｒｐ是保持抑制活性所必需的基团。水稻巯基蛋白酶抑制剂和丝氨酸蛋白酶抑制剂对稻瘟病菌丝体的生长均有抑制作用,但后者的抑制作用比前者更强,若将两种抑制剂混合使用,则对稻瘟病菌丝体的抑制作用非常强烈;当抑制剂加入量达７２μｇ时,即可产生明显的抑制作用。     

The native F0F1-inhibitor protein complex from beef heart mitochondria and its reconstitution in liposomes

A functional F₀F₁ ATP synthase that contains the endogenous inhibitor protein (F₀F₁I) was isolated by the use of two combined techniques [Adolfsen, R., McClung, J.A., and Moudrianakis, E. N. (1975).Biochemistry 14, 1727–1735; Dreyfus, G., Celis, H., and Ramirez, J. (1984).Anal. Biochem. 142, 215–220]. The preparation is composed of 18 subunits as judged by SDS-PAGE. A steady-state kinetic analysis of the latent ATP synthase complex at various concentrations of ATP showed aV _max of 1.28mol min^–1 mg^–1, whereas theV _max of the complex without the inhibitor was 8.3mol min^–1 mg^–1. In contrast, theK _m for Mg-ATP of F₀F₁ I was 148M, comparable to theK _m value of 142M of the F₀F₁ complex devoid of IF₁. The hydrolytic activity of the F₀F₁I increased severalfold by incubation at 60C at pH 6.8, reaching a maximal ATPase activity of 9.5mol min^–1 mg^–1; at pH 9.0 a rapid increase in the specific activity of hydrolysis was followed by a sharp drop in activity. The latent ATP synthase was reconstituted into liposomes by means of a column filtration method. The proteoliposomes showed ATP-Pi exchange activity which responded to phosphate concentration and was sensitive to energy transfer inhibitors like oligomycin and the uncouplerp-trifluoromethoxyphenylhydrazone.     

Cholinomimetics Increase Glutamate Outflow via an Action on the Corticostriatal Pathway: Implications for Alzheimer's Disease

Abstract: Physostigmine, the acetylcholinesterase inhibitor (0.3 mg/kg, i.m.), increased extracellular glutamate but not aspartate concentrations in the striatum of anaesthetised rats, determined using microdialysis and HPLC. The rise was both tetrodotoxin and calcium dependent. In contrast, neither physostigmine (10 µ M ) added to the perfusion fluid nor vehicle (injected intramuscularly) affected amino acid concentrations. To obtain evidence that the action of acetylcholine was to modulate positively cortical pyramidal neurone activity via the M₁ receptor, the selective M₁ agonist PD 142505-0028 (10 µ M ) was topically applied to the frontal cortex. Like physostigmine, PD 142505-0028 rapidly increased glutamate but not aspartate concentrations in the striatum. Moreover, the effect of intramuscular physostigmine was blocked by a topically applied M₁ antagonist. These new data add to our hypothesis that cholinomimetics increase pyramidal neurone function.     

A Protease Inhibitor of the Serpin Family Is a Major Protein in Carp Perimeningeal Fluid: I. Protein Purification and Characterization

Abstract: Two isoforms of a protease inhibitor of the serpin family (p62) have been purified from bighead carp perimeningeal fluid. Both isoforms migrate with an apparent molecular mass of 62 kDa on reducing and nonreducing sodium dodecyl sulfate-polyacrylamide gels. Both proteins inhibited the activities of bovine trypsin, bovine chymotrypsin, and porcine pancreatic elastase. They also formed complexes with these proteases that were resistant to sodium dodecyl sulfate treatment. p62 exists in the extracts of all tissues examined, including brain, head kidney, kidney, liver, muscle, ovary, pituitary, and spleen. It is also present in serum, ovarian fluid, and milt as well as perimeningeal fluid. The protease inhibitor is a glycoprotein, and its carbohydrate moiety could be removed by endoglycosidase F. Because p62 resembles mammalian α₁-antitrypsin in many aspects, it is likely a fish equivalent of α₁-antitrypsin.     

Thein vitro effect of theophylline on 17α, 20ß-dihydroxy-4-pregnen-3-one-induced germinal vesicle breakdown in the catfish,Clarias batrachus

The effects of theophylline (a phosphodiesterase inhibitor) and cAMP on 17α, 20ß-dihydroxy-4-pregnen-3-one-induced germinal vesicle breakdown was investigatedin vitro in catfish (Clarias batrachus) oocytes. Folliculated oocytes incubated with 17α, 20ß-dihydroxy-4-pregnen-3-one at the concentration of 1 μg/ml induced 93.2 ± 2.23% germinal vesicle breakdown. When the oocytes were prestimulated with 17α,20ß-dihydroxy-4-pregnen-3-one for 6 h and then treated with different concentrations of theophylline, there was a significant drop in the frequency of germinal vesicle breakdown at the concentrations 2.0, 1.5 and 1.0 mM. However, theophylline was found to be incapable of inhibiting germinal vesicle breakdown at its lowest concentration (0.5 inM). In the time course study, significant inhibition of germinal vesicle breakdown was recorded when 1 mM theophylline was added up to 30 h of 17α,20ß-dihydroxy-4-pregnen-3-one Stimulation but the inhibitory effect of theophylline gradually (time dependent manner) declined if the stimulatory time of 17α,20ß-dihydroxy-4-pregnen-3-one was increased. A similar inhibition of germinal vesicle breakdown was also recorded with various concentrations of cAMP. Except 0.5 mM, all the higher concentrations of cAMP significantly inhibited 17α,20ß-dihydroxy-4-pregnen-3-one induced germinal vesicle breakdown.     

Genetic control of PI and GC variants in the American Mink

Genetic polymorphism of the serum α-protease inhibitor (PI) and group-specific component (GC) in minks was revealed using one-dimensional polyacrylamide gel electrophoresis and immunoblotting. Two codominant alleles were identified at each of the two loci. The data ruled out the possibility of any linkage between the PI, GC and the coat colour gene Crystal ( Cr ).     

Determination of the disulfide bond pairings in human tissue factor pathway inhibitor purified fromEscherichia coli

The disulfide bond assignments of human alanyl tissue factor pathway inhibitor purified fromEscherichia coli have been determined. This inhibitor of the extrinsic blood coagulation pathway possesses three Kunitz-type inhibitor domains, each containing three disulfide bonds. The disulfide bond pairings in domains 1 and 3 were determined by amino acid sequencing and mass spectrometry of peptides derived from a thermolysin digest. However, thermolysin digestion did not cleave any peptide bonds within domain 2. The disulfide bond pairings in domain 2 were determined by isolating it from the thermolysin treatment and subsequently cleaving it with pepsin and trypsin into peptides which yielded the three disulfide bond pairings in this domain. These results demonstrate that the disulfide pairings in each of the three domains of human tissue factor pathway inhibitor purified fromEscherichia coli are homologous to each other and also to those in bovine pancreatic trypsin inhibitor.     

Tritium NMR studies of the human carbonic anhydrase I–benzenesulfonamide complex

Tritium NMR spectroscopy has been used to examine the complexformed by [4-³H]benzenesulfon-amide and human carbonicanhydrase I. The results show that in solution the inhibitor forms a 1:1complex with the enzyme. A 100-spin computational model of the system,constructed with reference to crystallographic results, was used tointerpret tritium relaxation behavior and ³H{¹H}NOEs. The analysis shows that the rate of dissociation of theenzyme–sulfonamide complex is 0.35 s^–1 and thatthe aromatic ring of the inhibitor undergoes rapid rotation while complexed.     

The medical utility of genomics data in neuropsychiatry: mutational genetics versus association genetics

The long anticipated ‘genetic revolution’ in neuropsychiatry has yet to have an impact on the practice of clinical medicine. Excitement in the 1980s over major genetic breakthroughs in schizophrenia and manic depression, for example, has been replaced in the late 1990s by the sobering realization that most common neuropsychiatric disorders are multifactorial. Despite considerable effort and resources, no ‘causative’ genetic variation has been identified that plays a definitive major role in any common neuropsychiatric disorder.