Kun Silprasit

Antibacterial activity of crude extracts of cyanobacteria Phormidium and Microcoleus species

Cyanobacteria is a group of prokaryotic organisms that occurred originally in the world. Thailand is well known as an area of a high biological diversity. In our study, eighteen cyanobacteria from the natural area were isolated and screened for their antibacterial activities. Of these, two isolated cyanobacteria (NCI1 and NCI4) with positive antibacterial activities were identified by their morphology and 16S rRNA sequences as Phormidium and Microcoleus group in family Oscillatoriaceae. The ethanol extracts of Phormidium sp. and Microcoleus sp. at various concentrations (0.2, 0.06, 0.03 and 0.015 g/ml) showed the antibacterial activity against Streptococcus enteritidis and Escherichia coli on the media. Each cyanobacterial extract showed more inhibition zones against S. enteritidis and E. coli by increasing their concentrations, especially those of Phormidium extracts. At the highest concentration 0.2 g/ml, the maximum diameter of inhibition were observed in the extracts of Phormidium sp. and Microcoleus sp., 11 and 10 mm against S. enteritidis, and 10 and 10 mm against E. coli, respectively. As results, the extracts of Phormidium sp. and Microcoleus sp. showed potentially interesting antibacterial activities against the two pathogens. Therefore, the two cyanobacteria may be useful in various applications and used as basic knowledge for further investigations.

Simple and rapid determination of the enzyme kinetics of HIV-1 reverse transcriptase and anti-HIV-1 agents by a fluorescence based method

Highly sensitive fluorescence dye, PicoGreen, can detect a small amount of polynucleotide duplex. The DNA/RNA duplex is a reaction product of HIV-1 reverse transcriptase (HIV-RT), which demonstrated that the HIV-1 RT kinetics and anti-HIV agent efficiency could be determined rapidly by a fluorometric method using PicoGreen dye. A small amount of dye is enough to test a minimal quantity of substrate to rapidly determine the enzyme kinetic properties K(m) and V(max). This method was applied to screen HIV-RT inhibitors. The enzymatic inhibition of 11 dipyridodiazepinone derivative inhibitors was compared. The IC(50) of all compounds was determined and it was found that two showed high inhibition efficiency. In addition, this method is applied for screening of many natural agents belonging to diverse sources. The proposed fluorometric method using PicoGreen dye is an easy, rapid, and sensitive detection method to determine HIV-RT activity, monitor enzyme kinetics and perform high-throughput screening of anti-HIV-1 RT agents.

Purification, crystallization and preliminary X-ray analysis of recombinant betaine aldehyde dehydrogenase 2 (OsBADH2), a protein involved in jasmine aroma, from Thai fragrant rice (Oryza sativa L.)

Fragrant rice (Oryza sativa L.) betaine aldehyde dehydrogenase 2 (OsBADH2) is a key enzyme in the synthesis of fragrance aroma compounds. The extremely low activity of OsBADH2 in catalyzing the oxidation of acetaldehyde is believed to be crucial for the accumulation of the volatile compound 2-acetyl-1-pyrroline (2AP) in many scented plants, including fragrant rice. Recombinant fragrant rice OsBADH2 was expressed in Escherichia coli as an N-terminal hexahistidine fusion protein, purified using Ni Sepharose affinity chromatography and crystallized using the microbatch method. Initial crystals were obtained within 24 h using 0.1 M Tris pH 8.5 with 30%(w/v) PEG 4000 and 0.2 M magnesium chloride as the precipitating agent at 291 K. Crystal quality was improved when the enzyme was cocrystallized with NAD(+). Improved crystals were grown in 0.1 M HEPES pH 7.4, 24%(w/v) PEG 4000 and 0.2 M ammonium chloride and diffracted to beyond 2.95 Å resolution after being cooled in a stream of N(2) immediately prior to X-ray diffraction experiments. The crystals belonged to space group C222(1), with unit-cell parameters a = 66.03, b = 183.94, c = 172.28 Å. An initial molecular-replacement solution has been obtained and refinement is in progress.

The partial mitochondrial sequence of the Old World stingless bee, Tetragonula pagdeni

1Faculty of Health Science, Srinakharinwirot University Ongkharak, Nakhon-Nayok 26120, Thailand 2Department of Biochemistry, Faculty of Science, Chulalongkorn University, Bangkok 10330, Thailand 3Faculty of Environmental Culture and Ecotourism, Srinakharinwirot University, Nakhon-Nayok 26120, Thailand 4Aquatic Molecular Genetics and Biotechnology, National Science Center for Genetic Engineering and Biotechnology (NSTDA), 113 Paholyothin Road, Klong 1, Klong Luang, Pathum Thani 12120, Thailand