Identification of the zinc binding ligands and the catalytic residue in human aspartoacylase, an enzyme involved in Canavan disease

Canavan disease is an autosomal-recessive neurodegenerative disorder caused by a lack of aspartoacylase, the enzyme that degrades N-acetylaspartate (NAA) into acetate and aspartate. With a view to studying the mechanisms underlying the action of human aspartoacylase (hASP), this enzyme was expressed in a heterologous Escherichia coli system and characterized. The recombinant protein was found to have a molecular weight of 36 kDa and kinetic constants K(m) and k(cat) of 0.20 +/- 0.03 mM and 14.22 +/- 0.48 s(-1), respectively. Sequence alignment showed that this enzyme belongs to the carboxypeptidase metalloprotein family having the conserved motif H(21)xxE(24)(91aa)H(116). We further investigated the active site of hASP by performing modelling studies and site-directed mutagenesis. His21, Glu24 and His116 were identified here for the first time as the residues involved in the zinc-binding process. In addition, mutations involving the Glu178Gln and Glu178Asp residues resulted in the loss of enzyme activity. The finding that wild-type and Glu178Asp have the same K(m) but different k(cat) values confirms the idea that the carboxylate group contributes importantly to the enzymatic activity of aspartoacylase.     

紫色色杆菌感染临床特症及耐药谱分析

目的 研究紫色色杆菌感染的临床特征与病原体的生物学特性以及耐药分析,以提高临床对紫色色杆菌感染的诊断和治疗.方法 将患者的血液、浓液标本进行培养,分离并把分离获得的菌株在VITEK-32微生物鉴定仪中进行种的鉴定及药物敏感性试验,用小白鼠进行该病原体的毒力试验.结果 12种抗菌药物中舒普深、亚胺培南、特治星（派拉西林/他唑巴坦）、马斯平（头孢吡肟）及喹喏酮类抗菌药物对紫色色杆菌均有较强的抗菌作用,而氨苄西林、头孢唑啉、庆大霉素、妥布霉素、氨曲南、头孢曲松和头孢他啶等抗菌药物抗菌效果不佳.毒力试验结果表明该菌有较强毒力.结论 紫色色杆菌是一种革兰阴性杆菌,毒力强.可引起感染部位脓肿,并可侵入体内各脏器及血液引发脏器的脓肿,脓毒血症是该菌感染的临床特症,重者可引起各脏器功能衰退,中毒性休克甚至死亡.紫色色杆菌感染病情急,进展快,死亡率高,因此早发现,及早有效性治疗极为重要.     

群体感应系统对紫色杆菌CV31532积累PHA的影响

紫色杆菌CV31532中群体感应系统产生的信号分子C6-HSL是由cviI基因编码合成的。在限氮条件下,以D-葡萄糖酸钠、果糖、葡萄糖为唯一碳源时,紫色杆菌CV31532均产生聚-3-羟基丁酸,且以D-葡萄糖酸钠为唯一碳源时积累量最高。利用气相色谱分析发现,紫色杆菌CV31532培养48 h的PHA积累量显著高于其群体感应合成酶突变株CV026 PHA的积累量;通过薄层层析分析发现紫色杆菌CV31532合成信号分子的量在48 h显著提高;外源添加C6-HSL信号分子可显著提高突变体CV026 PHA的积累量。由此推测,紫色杆菌群体感应系统参与调控胞内PHA的合成。     

具有群体感应抑制活性的海洋细菌的筛选

【目的】从海洋环境中筛选出能有效抑制细菌群体感应的活性菌株,为以致病菌群体感应为靶点的新型疗法提供新的天然产物资源。【方法】以紫色杆菌(Chromobacteriumviolaceum)为报告菌,采用滤纸片法和双层软琼脂法相结合的筛选模型进行群体感应抑制活性菌的筛选。【结果】通过对美国圣璜岛海域海绵中分离出来的272株海洋细菌群体感应抑制活性的筛选,得到了具有抑制紫色杆菌素产生的细菌51株,其中74号菌株抑制效果最好,具有进一步研究的价值。【结论】海洋细菌中有很多具有抑制细菌群体感应效应的菌株,是天然群体感应抑制剂的潜在来源。     

Syngenic Divisions of the Cellular Slime Mold Polysphondylium violaceum*

SYNOPSIS. Mating type analysis of new isolates of Polysphondylium violaceum supports the subdivision of this morphological species into two reproductively isolated breeding groups or syngens. Representatives of both syngens have been identified in soil samples taken from widely separated geographical locations. Intersyngenic cross reactions have been observed in some stocks.     

Pathogen hijacks programmed cell death signaling by arginine ADPR-deacylization of caspases

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