Soriano Margarita, Blanco Ana, Dı Az Pilar, Pastor F I Javier
Department of Microbiology, Faculty of Biology, University of Barcelona, Avinguda Diagonal 645, 08028 Barcelona, Spain1.
Microbiology (Reading). 2000 Jan;146 ( Pt 1):89-95. doi: 10.1099/00221287-146-1-89.
The gene pelA encoding a pectate lyase from the strain Bacillus sp. BP-23 was cloned and expressed in Escherichia coli. The nucleotide sequence of a 1214 bp DNA fragment containing pelA gene was determined, revealing an ORF of 666 nucleotides that encoded a protein of 23233 Da. The deduced amino acid sequence of the encoded enzyme showed homology to pectate lyases A, B, C and D from Fusarium solani, Pel-3 and PelB from Erwinia carotovora and Pell from Erwinia chrysanthemi. Homology was also found to the protein deduced from the Bacillus subtilis yvpA gene, the function of which is unknown. The heterologous expressed enzyme depolymerized polygalacturonate and pectins of methyl esterification degree from 22 to 89%, and exhibited similar activity on polygalacturonate and on 89% esterified citrus pectin. Optimum temperature and pH for enzymic activity were 50 degrees C and pH 10, respectively. Ca2+ was required for activity on pectic substrates, while the enzyme was strongly inhibited by Ba2+.
克隆了芽孢杆菌属BP - 23菌株中编码果胶酸裂解酶的基因pelA，并在大肠杆菌中进行表达。测定了包含pelA基因的1214 bp DNA片段的核苷酸序列，发现一个666个核苷酸的开放阅读框，编码一个23233 Da的蛋白质。所编码酶的推导氨基酸序列与茄腐镰刀菌的果胶酸裂解酶A、B、C和D、胡萝卜软腐欧文氏菌的Pel - 3和PelB以及菊欧文氏菌的Pell具有同源性。还发现与枯草芽孢杆菌yvpA基因推导的蛋白质具有同源性，其功能未知。异源表达的酶能使聚半乳糖醛酸和甲酯化程度为22%至89%的果胶解聚，并且对聚半乳糖醛酸和89%酯化的柑橘果胶表现出相似的活性。酶活性的最适温度和pH分别为50℃和pH 10。在果胶底物上的活性需要Ca2 +，而该酶受到Ba2 +的强烈抑制。
