Controlled potential enzymology of methyl transfer reactions involved in acetyl-CoA synthesis by CO dehydrogenase and the corrinoid/iron-sulfur protein from Clostridium thermoaceticum

Many anaerobic bacteria fix CO2 via the Wood pathway of acetyl-CoA synthesis. Carbon monoxide dehydrogenase (CODH), also called acetyl-CoA synthase, accepts the methyl group from the methylated corrinoid/iron-sulfur protein (C/Fe-SP), binds a carbonyl group from CO, CO2, or the carboxyl of pyruvate, and binds coenzyme A. Then CODH catalyzes the synthesis of acetyl-CoA from these enzyme-bound groups. Here, we have characterized the methyl transfer steps involved in acetyl-CoA synthesis. We have studied the reactions leading to methylation of CODH by methyl iodide and shown an absolute requirement of the C/Fe-SP in this reaction. In addition, we have discovered and partly characterized two previously unknown exchange reactions catalyzed by CODH: between the methylated C/Fe-SP and methylated CODH and between methylated CODH and the methyl moiety of acetyl-CoA. We have performed these two exchange reactions, methylation of the C/Fe-SP, and methylation of CODH at controlled potentials. The rates of all these reactions except the exchange between methylated C/Fe-SP and methylated CODH are accelerated (from 1 to 2 orders of magnitude) when run at low potentials. Our results provide strong evidence for a nucleophilic redox-active metal center on CODH as the initial acceptor of the methyl group from the methylated C/Fe-SP. This metal center also is proposed to be involved in the cleavage of acetyl-CoA in the reverse reaction.     

Effects of chloramine on Bacillus subtilis deoxyribonucleic acid.

The lesions induced in Bacillus subtilis deoxyribonucleic acid (DNA) after treating bacterial cells (in vivo) and bacterial DNA (in vitro) with chloramine were studied biologically and physically. Single-strand breaks and a few double-strand scissions (at higher chloramine doses) accompanied loss of DNA-transforming activity in both kinds of treatments. Chloramine was about three times more efficient in vitro than in vivo in inducing DNA single-strand breaks. DNA was slowly chlorinated; the subsequent efficiency of producing DNA breaks was high. Chlorination of cells also reduced activity of endonucleases in cells; however, chlorinated DNA of both treatments was sensitized to cleavage by endonucleases. The procedure of extracting DNA from cells treated with chloramine induced further DNA degradation. Both treatments introduced a small fraction of alkali-sensitive lesions in DNA. DNA chlorinated in vitro showed further reduction in transforming activity as well as further degradation after incubation at 50 C for 5 h whereas DNA extracted from chloramine-treated cells did not show such a heat sensitivity.     

l-Ornithine:2-Oxoacid Aminotransferase from Squash (Cucurbita pepo, L.) Cotyledons: Purification and Properties

ORNITHINE: 2-oxoacid aminotransferase (EC 2.6.1.13) has been purified over 400-fold with a total recovery of 14% from acetone powders of cotyledons of germinating squash (Cucurbita pepo, L.) seedlings. The pH optimum of the transamination between l-ornithine and alpha-ketoglutarate is 8 and the Michaelis constants are 4.7 mm and 6.3 mm, respectively. The enzyme has a molecular weight of 48,000 as determined by gel filtration. The reaction is essentially specific for alpha-ketoglutarate as the amino group acceptor. The enzyme is inhibited very strongly by hydroxylamine, and less severely by NaCN and isonicotinylhydrazide. No inhibition is observed in the presence of 10 mml-cysteine. The energy of activation is 7.6 kcal/mole. The stability of the enzyme preparation is enhanced by the presence of dithioerythritol and glycerol. The enzyme activity of the most purified fraction is stimulated 30% by the addition of pyridoxal phosphate; however, the evidence for the unequivocal involvement of pyridoxal phosphate was inconclusive.     

Assessing the risk of resistance to cationic biocides incorporating realism-based and biophysical approaches

The control of microorganisms is a key objective in disease prevention and in medical, industrial, domestic, and food-production environments. Whilst the effectiveness of biocides in these contexts is well-evidenced, debate continues about the resistance risks associated with their use. This has driven an increased regulatory burden, which in turn could result in a reduction of both the deployment of current biocides and the development of new compounds and formulas. Efforts to balance risk and benefit are therefore of critical importance and should be underpinned by realistic methods and a multi-disciplinary approach, and through objective and critical analyses of the literature. The current literature on this topic can be difficult to navigate. Much of the evidence for potential issues of resistance generation by biocides is based on either correlation analysis of isolated bacteria, where reports of treatment failure are generally uncommon, or laboratory studies that do not necessarily represent real biocide applications. This is complicated by inconsistencies in the definition of the term resistance. Similar uncertainties also apply to cross-resistance between biocides and antibiotics. Risk assessment studies that can better inform practice are required. The resulting knowledge can be utilised by multiple stakeholders including those tasked with new product development, regulatory authorities, clinical practitioners, and the public. This review considers current evidence for resistance and cross-resistance and outlines efforts to increase realism in risk assessment. This is done in the background of the discussion of the mode of application of biocides and the demonstrable benefits as well as the potential risks.     

合成己酸乙酯脂肪酶产生菌的筛选及产酶条件

从２７株脂肪酶产生菌中筛选到能由乙醇和己酸合成己酸乙酯的菌株８株。其中Ｒｈｉｚｏｐｕｓｓｐ．Ｈ－３菌株脂肪酶活力为５０－６０ｕ／ｍｌ,全细胞在有机溶剂中的酯化率可达己酸的９１％。Ｈ３产酶的最适碳源为淀粉或葡萄糖。６％黄豆饼粉加４％蛋白陈复合氮源有利于酶活力的增加。     

缺氧预适应研究的进展与展望

缺氧预适应这一生物进化上的内源性细胞保护机制,可被机体、器官、组织和细胞的重复缺氧暴露所激发。缺氧预适应的效应已由对重复缺氧局部／原位器官组织的保护（局部／原位缺氧预适应）发展到既保护远隔的各种异位器官组织（远程／异位缺氧预适应）又抗御其它种种非缺氧性应激（交叉／多能缺氧预适应）。在现有进展的基础上,缺氧预适应研究以及其可操作性和可应用性将有更大的发展空间。