Lignolytic Enzyme Activity of Isolated Bacteria from Termite (Coptotermes Sp.) and Milkfish (Chanos chanos Forsskal, 1775) Guts

Bacteria BSR 2, Pseudomonas alcaligenes (BSR 3), Brevibacillus parabrevis (BSR 8), Brevibacillus sp. (BSR 9), isolated from termite gut and Bacillus licheniformis (BSA B1) isolated from milkfish gut have been known to possess celluloytic activity. However, their lignolytic ability has not been known. This study aimed to determine the lignolytic ability of bacteria isolated from termit (Coptotermes sp.) and milkfish (Chanos chanos Forsskal, 1775) guts and their enzymes characterization. The qualitative test was done through the spot test method, while quantitative assay was performed spectrophotometrically at 335 nm to calculate vanillin concentration. The isolates were grown in Lignin Mineral Medium, then the optical density (OD620) were measured every 24 hours for 5 days using spectrophotometer to determine their growth profile and the best isolation time of the lignolytic enzyme. Based on results, the best lignolytic enzyme isolation time for strains Bacillus licheniformis (BSA B1) and BSR 2 were 5 days, yielding lignolytic enzyme activity of 0.961 ± 0.168 U/mg and 2.176 ± 0.088 U/mg respectively,  while strains Pseudomonas alcaligenes (BSR 3), Brevibacillus parabrevis (BSR 8), and Brevibacillus sp. (BSR 9) were 4 days, yielding of 1.206 ± 0.045 U/mg, 1.162 ± 0.191 U/mg, and 0.896 ± 0.108 U/mg, respectively. The strain BSR 2 showed the highest lignolytic activity compared to other strains. The optimum temperature for lignolytic enzyme activity of BSR 2 was 30 ℃ and the optimum pH was 7. The lignolytic enzyme activity showed that these bacterial isolates can be a chance to be used as new alternative lignolytic enzyme source in commercial bioconversion process.

Eksplorasi Mikroba Penghasil Enzim-enzim Hidrolitik Di Kawasan Taman Nasional Lore Lindu Sulawesi Tengah

Lore Lindu National Park (TNLL) is an area that flora, fauna and microbes, the diversity of microbes producing hydrolytic enzyme. explore the hydrolytic enzyme producing microbes in Indonesia. This study was aimed to obtain bacterial isolates were able to produce hydrolytic enzyme and characteristics. Isolation in the microbiology laboratory. Isolation by a spread plate. Isolates in the selection hydrolytic enzyme producing selective media. Measurement of the activity of the enzyme with hydrolytic index. The results were thirteen isolates clearing zone test , 2 protease enzyme bacterial isolates, 1 lipase enzyme bacterial isolates, 6 amylase enzyme bacterial isolates, while 4 cellulase enzyme bacterial isolates. Examination of Amylase enzyme activity was done using DNS method. L10T3 showed that the bacterial isolate optimum activity at pH 7 and at a temperature of 300C with an activity of 0.040 U / mL and 0.029 U / mL. Key word: Lore Lindu National Park, hydrolytic enzyme. isolation of bacteria

ISOLATION AND PURIFICATION OF LACTASE FROM LACTOBACILLUS ACIDOPHILUS

The enzyme β-galactosidase (β-D-galactoside galactohydrolase, EC 3.2.1.23) commonly known as lactase, has important applications in the dairy industry. From culture of strain Lactobacillus acidophilus having high and stable β-galactosidase activity, the study of crude enzyme isolation were carried out by ultrasonical extraction and precipitation by neutral salts and organic solvents. Best precipitant was isopropanol with enzyme recovery 89,93%, and enzyme purity increased 4,5 folds. Further β-galactosidase purification was carried out using gel permeation chromatography on Ultrahydrogel 250 to increase purity in 14,3 folds. The molecular weight of β-galactosidase was 40 kDa. The purified enzyme had optimum activity at 40oC, pH 7 – 7,5 and kinetic parameters of Vmax and Km were 2,3 μmol/min and 0,73 mM.

Continuous assays of glutaminyl cyclase: from development to application

Glutaminyl cyclase (QC, EC 2.3.2.5) catalyses the formation of pyroglutamyl residues from glutamine at the N-terminus of peptides and proteins. In previously applied assays, QC activity was determined by either analysing the products formed using HPLC coupled with photometric or fluorometric detection, radioimmunoassay, or by detecting the release of ammonia spectrophotometrically. Although these methods are sensitive, they are all discontinuous and therefore time-consuming and laborious. To conduct a detailed kinetic investigation of QC catalysis, we developed coupled continuous assays suitable for microplates which allow now convenient determination of QC activity. The methods either use pyroglutamyl aminopeptidase or glutamate dehydrogenase as auxiliary enzymes, which results in the liberation of chromophores or fluorophores such aspNA, AMC,βNA or in the conversion of the chromophore NADH/H+into NAD+, respectively.The assays were applied in various enzyme isolation and characterisation studies, using crude protein solutions as well as purified enzyme in pH-dependence, substrate and inhibitor specificity investigations. Depending on the respective analytical task, both assays complement each other. Therefore, different enzymatic properties could be explored in more detail. Since the employed strategy of assay development could be of interest also for the analysis of other enzymes, the methods are described here in a comprehensive manner.