Specific and sensitive plate assay for bacterial lipases

Rathi

Biotech and App Biochem

et al. 2003

162

Lipases are versatile biocatalysts that can perform innumerable different reactions. Their enantio-, chemo- and stereo-selective nature makes them an important tool in the area of organic synthesis. Unlike other hydrolases that work in aqueous phase, lipases are unique as they act at the oil/water interface. Besides being lipolytic, lipases also possess esterolytic activity and thus have a wide substrate range. Hence, the lipase assay protocols hold a significant position in the field of lipase research. Lipase activity can be estimated using a wide range of assay protocols that differ in terms of their basic principle, substrate selectivity, sensitivity and applicability. As the value of these enzymes continues to grow and new markets are exploited, development of new or improved enzymes will be a key element in the emerging realm of biotechnology. Hence, development of faster and simpler protocols incorporating newer and more specific substrates is the need of the hour. In this endeavour, methods that could be adopted for molecular screening occupy an important position. Here, an overview of the lipase assay protocols is presented with emphasis on the assays that can be adopted for the molecular screening of these biocatalysts.

Section: Estimation Of Fatty Acids Releasedmentioning

confidence: 99%

Lipase assays for conventional and molecular screening: an overview

Rathi

Biotech and App Biochem

et al. 2003

162

FEMS Microbiology Letters

“…Presence of lipolytic activity in B. megaterium 370 was analysed using Rhodamine B-supplemented plates [11], containing tributyrin or olive oil. The strain showed good performance on lipid-based substrate hydrolysis, as con¢rmed by £uorescence emission from Rhodamine B and appearance of hydrolysis areas around the colonies (not shown).…”

Section: Analysis Of B Megaterium 370 Lipolytic Systemmentioning

confidence: 99%

Analysis of Bacillus megaterium lipolytic system and cloning of LipA, a novel subfamily I.4 bacterial lipase

Ruiz¹

2002

The lipolytic system of Bacillus megaterium 370 was investigated, showing the existence of at least two secreted lipases and a cell-bound esterase. A gene coding for an extracellular lipase was isolated and cloned in Escherichia coli. The cloned enzyme displayed high activity on short to medium chain length (C 4^C8 ) substrates, and poor activity on C 18 substrates. On the basis of amino acid sequence homology, the cloned lipase was classified into subfamily I.4 of bacterial lipases.

FEMS Microbiology Ecology

“…Standard methods were used for Gram staining and testing for aminopeptidase, oxidation fermentation and gas formation, activities of oxidase and catalase and gelatin hydrolysis [20]. Secretion of lipase was tested on rhodamine B medium [21]. Partial 16S RNA gene sequencing and sequence alignments were performed by DSMZ [22,23].…”

Section: Phenotypic Tests and Sequencing Of 16s Rrna Genesmentioning

confidence: 99%

Interactions between bacterial populations during degradation of a lubricant base oil

Sonderkamp

2001

The composition of microbial communities degrading trimethylolpropaneoleate, a synthetic fatty acid ester, was studied under conditions similar to a frequently used degradation test. A combination of conventional phenotypic tests and molecular biological methods, internal rRNA gene spacer amplification and randomly amplified DNA assay, was used for strain monitoring. Few bacterial species were selected from the large diversity of the inocula during degradation in the batch cultures. Growth depended on activity of extracellular lipases in the bacterial community. However, the enzyme producers made up only a minority. The dominating bacteria were not able to hydrolyze the synthetic ester in pure culture. ß