Alkaliphilic Bacillus species and an alkaline active lipolytic enzyme

Detta är en avhandling från Biotechnology, Lund University

Sammanfattning: The Great Rift Valley running through Eastern Africa contains a large number of soda lakes. These unique lakes constitute among the most stable alkaline environments with a dense and varied population of microorganisms. Such microorganisms are invariable alkaliphiles and are considered to be a promising source of enzymes with unique features. Some enzymes produced by alkaliphilic microorganisms have found their way to industrial applications.Phylogenetic analysis and morphological and physiological characteristics of the isolates from a screening for lipase-producing microorganisms from a Kenyan alkaline soda lake showed some of them to be closely related to the Pseudomonas and Halomonas cluster sharing high similarity profile with Halomonas desiderata. The remaining were Gram-positive, closely related to the Bacillus cluster, and were grouped with Bacillus halodurans, Bacillus alcalophilus and Bacillus licheniformis, respectively. A new alkaliphilic Bacillus sp. was identified during the screening and based on detailed studies we proposed a novel alkaliphilic, halotolerant species of the genus Bacillus with the name Bacillus bogoriensis.Among the alkaliphilic Bacillus species isolated, Bacillus halodurans was selected due to its remarkable enzymes studied so far. A lipolytic enzyme was cloned and overexpressed in Escherichia coli. Production of the enzyme was mainly in the form of inclusion bodies from which recovery of the active enzyme was achieved. Since E. coli proved not to be a suitable host for production of soluble enzyme, Pichia pastoris was tested as an alternative host and the production of the heterologous enzyme in a soluble form was also studied. The enzyme was characterized as a phospholipase, also displaying hydrolysis of soluble and insoluble fatty substrates, and esterification of fatty acids.