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Articles by issue : Vol 9, No 2 (2015): June 2015
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Articles
Expression and Purification of PhoR Sensor-Domain Histidine Kinase of Mycobacterium tuberculosis in Escherichia coli

GIRI-RACHMAN, ERNAWATI ARIFIN, PRATAMA, FENRYCO, AJI, OKTIRA ROKA, PATRIATI, ARUM, IHSANAWATI, IHSANAWATI, MOEIS, MAELITA RAMDANI, PUTRA, EDY GIRI-RACHMAN

Microbiology Indonesia Vol 9, No 2 (2015): June 2015
Publisher : Indonesian Society for microbiology

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Abstract

Globally, tuberculosis (TB) remains a leading cause of death. The emergence of multidrug-resistant strains (MDR-TB) and extensively drug-resistant strains (XDR-TB) has fuelled the discovery for novel drugs and drug targets for its successful and better treatment. One of the potential candidates for drug target is PhoR sensory protein histidine kinase, a part of the Two Component System (TCS) PhoP/PhoR in Mycobacterium tuberculosis (Mtb). This protein system was known for its role on regulating hundred of Mtb virulence factors, from genes for cell wall and lypid synthesis to genes for adaptation in human leukocyte and hypoxia response. Previous studies have successfully characterized, isolated, and cloned the putative sensory domain of PhoR protein gene into pRSET vector expression system. In this study, Escherichia coli was transformed with pRSET-SensPhoR and cultivated at 37oC under IPTG induction to express PhoR sensor-domain protein. Most of the proteins were overexpressed in the form of inclusion bodies.  Subsequent protein purification in Ni-NTA system under refolding condition on urea gradient was performed to isolate PhoR sensor-domain protein in soluble form. Arginine was supplemented in purified protein solution to prevent aggregation during long term storage.  While highly purified protein was acquired, small angle X-ray scattering (SAXS) analysis was conducted to obtain 3-dimensional (3D) protein structures in solution.    doi:10.5454/mi.9.2.1 

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Rusmana, Iman ( 1. Institut Pertanian Bogor 2. Indonesian Society for Microbiology )

Microbiology Indonesia Vol 9, No 2 (2015): June 2015
Publisher : Indonesian Society for microbiology

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Abstract

Genetic Profiles of Escherichia coli Isolated from Indonesian Tempeh Based on Enterobacterial Repetitive Intergenic Consensus- Polymerase Chain Reaction (ERIC-PCR)

A’YUN, QURROTA, SUWANTO, ANTONIUS, BARUS, TATI

Microbiology Indonesia Vol 9, No 2 (2015): June 2015
Publisher : Indonesian Society for microbiology

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Abstract

Tempeh is a famous Indonesian fermented food derived from soybeans inoculated with Rhizopus sp. Tempeh production varies depend on the producers and often conducted in an uncontrolled condition. This condition could lead to the growth of Escherichia coli which is known as bacterial indicators of environmental hygiene. Some strains of E. coli could induce diarrhea, acute gastroenteritis or gastrointestinal tract infections. The aim of this study was to compare genetic diversity of E. coli isolates from tempeh with medical isolates employing ERIC-PCR method. In this study, 63f and 1387r primers were used to amplify 16S rRNA genes, and ERIC 1R and ERIC 2 primers were used for ERIC-PCR analysis. Tempeh samples were obtained from four producers in Bogor. Thirty-three isolates of E. coli were successfully isolated from tempeh samples produced by only two producers, we could not obtain E. coli isolates from the other two producers. In addition, the same tempeh samples could carry different genotypes of E. coli. On the other hand, the same genotypes could be found in different tempeh samples. Based on phylogenetic tree analysis, E. coli from tempeh could be separated from medical isolates. We showed that E. coli isolates derived from tempeh were genetically different from those of medical or pathogenic isolates.

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Rusmana, Iman ( 1. Institut Pertanian Bogor 2. Indonesian Society for Microbiology )

Microbiology Indonesia Vol 9, No 2 (2015): June 2015
Publisher : Indonesian Society for microbiology

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Abstract

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Rusmana, Iman ( 1. Institut Pertanian Bogor 2. Indonesian Society for Microbiology )

Microbiology Indonesia Vol 9, No 2 (2015): June 2015
Publisher : Indonesian Society for microbiology

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Abstract

Chemical Constituen from an Endophytic Fungus Aspergillus sp (SbD5) Isolated from Sambiloto (Andrographis paniculata Nees)

ELFITA, ELFITA, MUNAWAR, MUNAWAR, MUHARNI, MUHARNI, IVANTRI, IHSAN

Microbiology Indonesia Vol 9, No 2 (2015): June 2015
Publisher : Indonesian Society for microbiology

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Abstract

Medicinal plants and their endophytes are important resources for discovery of natural products. Endophytic fungi isolated from medicinal plants more likely exibit pharmaceutical potentials. In the present study, an endophytic fungus Aspergillus sp (SbD5) was isolated from leaves of sambiloto (Andrographis paniculata). The fungus isolate was cultivated in Potato Dextrose Broth (PDB) medium for 7 weeks in static, then extracted with ethyl acetate followed by Thin Layer Chromatography (TLC) test. The results displayed three major spots. Ethyl acetate extract was further separated by column chromatography and recrystallization to obtain three pure compounds. Their structures were determined on the basic of spectroscopic analysis. The compounds is one new benzochromen derivative, 1-(3,8-dihydroxy-4,6,6-trimethyl-6H- benzochromen-2-yloxy)propan-2-one (1), together with two known compounds 5-hydroxy-4-(hydroxymethyl)-2H-pyran-2-one (2) and (5-hydroxy-2-oxo-2H-pyran-4-yl)methyl acetate (3). The antibacterial activities of the compounds were tested using the disk diffusion method against Staphylococcus aureus (ATCC 25923), Escherichia coli (ATCC 25922), Shigella dysenteriae and Salmonella typhi.  Compound 1 has the highest antibacterial activity followed by compound 2 and 3.

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Rusmana, Iman ( 1. Institut Pertanian Bogor 2. Indonesian Society for Microbiology )

Microbiology Indonesia Vol 9, No 2 (2015): June 2015
Publisher : Indonesian Society for microbiology

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Abstract

Inhibitory activity of Lactobacillus plantarum U10 isolated from Tempoyak (fermented durian) Made in Indonesia against Salmonella typhi

SOGANDI, SOGANDI, MUSTOPA, APON ZAENAL, ARTIKA, I MADE, BUDIARTO, BUGI RATNO

Microbiology Indonesia Vol 9, No 2 (2015): June 2015
Publisher : Indonesian Society for microbiology

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Abstract

Lactobacillus plantarum U10 produced bacteriocin U10 which was isolated from a traditionally fermented food “tempoyak” from Sumatera Island in Indonesia. Production of the bacteriocins started at early exponential phase and reached maximum level at early stationary phase. Furthermore, plantaricins U10 was purified by ammonium sulphate precipitation followed by gel filtration chromatography. L. plantarum U10 produced two bacteriocins with a molecular mass of approximately 4.5 and 9.8 kDa by SDS-PAGE.  The mode of action of plantaricins U10 was identified as bactericidal agents against Salmonella typhi ATCC25241 as proven by CFU counting and SEM micrographs that showed differences in cell structures between treated cells and the non-treated control. SEM examination also confirmed structural destruction of membrane cells integrity and considerable morphological alteration of S.typhi.

Medium Optimization for Penicillin Acylase (PAc) Production by Recombinant B. megaterium MS941 Containing pac Gene from B. thuringiensis BGSC BD1 Using Response Surface Methodology

PUTRI, FENTRI PARAMITHA, NURHASANAH, ASTUTIATI, NURHAYATI, NIKNIK, HELIANTI, IS, SYAMSU, KHASWAR

Microbiology Indonesia Vol 9, No 2 (2015): June 2015
Publisher : Indonesian Society for microbiology

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Abstract

Penicillin G acylase (PAc) hydrolyses of the amide bond of benzylpenicillin (Pen-G) releasing PAA and 6-APA, key intermediate in the production of various semisynthetic penicillins. In this study, we optimised the production medium of PAc by RSM using two variables (xylose as inducer and CaCl2 as divalent cations) to obtain the optimum PAc specific activity from Bacillus megaterium btpacBD1. For this purpose, combinations of five different xylose concentrations (0.13 – 0.87 %) and five different CaCl2 concentrations (0.64 – 4.36 mM) were analysed, in a total of 22 experiments. CCD used for the analysis showed that in shake flask cultivations, xylose and CaCl2 showed significant effects on PAc volumetric activity and the quadratic model was in good agreement with the experimental results (R2= 0.86 (p-value < 0.0001)). The maximum specific activity (130.669 ± 50.241 units mg protein-1) was reached when xylose and CaCl2 concentrations were 0.49% and 2.4 mM, respectively, and medium pH was around 7. Under such conditions, the activity of PAc and protein concentration achieved were 1.318 ± 0.406 units mL-1 and  0.0101 ± 0.01 mg mL-1. The shake flask validation experiments demonstrated that with such medium composition the volumetric activity, protein concentration and specific activity achieved were 1.294 ± 0.171 units mL-1, 0.0102 ± 0.0003 mg mL-1 and 125.91 ± 13.309 units mg-1, respectively. When the optimum medium composition was applied in 10 L bioreactor, the optimum volumetric activity (2.0687 ± 0.0820 units mL-1) and protein concentration (0.0078 ± 0.0008 mg mL-1) were achieved 48 h after the start of the cultivation. However, the optimum PAc specific acivity (1260.52  ± 27.5711 units mg protein-1) was achieved 18 h after the start of the cultivation.

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Rusmana, Iman ( 1. Institut Pertanian Bogor 2. Indonesian Society for Microbiology )

Microbiology Indonesia Vol 9, No 2 (2015): June 2015
Publisher : Indonesian Society for microbiology

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Abstract