Pub Date : 2018-12-31DOI: 10.14203/ANN.BOGOR.2018.V22.N2.47-56
A. Solikhin, A. Z. Mustopa, S. Suharsono, W. Putranto
Lactobacillus casei WSP-derived an aspartic protease was sequentially purified by using chromatography gel filtration sephadex G-50. It resulted in a 22.81-fold increase of specific activity (51.5 U/mg) with a final yield of 1.9%. The estimated molecular weight of the purified enzyme was 37 kDa and showed gelatinolytic activity in zymogram assay. The enzyme exhibited optimum activity at 40ºC and pH 6 with casein as the substrate. Enzyme activity was significantly inhibited by pepstatin A (0.5 mM and 1 mM), confirming that this enzyme is a group of aspartic proteases, while other inhibitors such as EDTA, PMSF and iodoacetic acid showed no inhibition effect on the activity of enzyme. The addition of metal ion to the enzyme decreased enzyme activity, indicating the proteolytic enzyme was metal ion- dependent. Denaturant such as DDT tended to increase caseinolytic activity. Furthermore, this enzyme was capable of generating the new peptides from skimmed milk with the size 8 kDa, 10 kDa and 15 kDa. These peptides have potential as antibacterial and antioxidant agents.
采用sephadex G-50层析凝胶过滤法对干酪乳杆菌wsp衍生的天冬氨酸蛋白酶进行了纯化。比活性提高22.81倍(51.5 U/mg)，最终产率为1.9%。酶谱分析表明，纯化酶的分子量为37 kDa，具有溶胶活性。以酪蛋白为底物，在40℃和pH 6条件下酶活性最佳。酶活性被pepstatin A (0.5 mM和1 mM)显著抑制，证实该酶是一组天冬氨酸蛋白酶，而其他抑制剂如EDTA、PMSF和碘乙酸对酶活性无抑制作用。金属离子的加入降低了酶的活性，表明该蛋白水解酶是金属离子依赖性的。变性剂如滴滴涕倾向于增加溶酪蛋白活性。此外，该酶还能从脱脂牛奶中生成8kda、10kda和15kda的新肽。这些肽具有抗菌和抗氧化剂的潜力。
{"title":"Partial Purification, Characterization, and Application of Extracellular Aspartic Protease from Lactobacillus casei WSP in Producing the Bioactive Peptides with Antibacterial and Antioxidant Activity","authors":"A. Solikhin, A. Z. Mustopa, S. Suharsono, W. Putranto","doi":"10.14203/ANN.BOGOR.2018.V22.N2.47-56","DOIUrl":"https://doi.org/10.14203/ANN.BOGOR.2018.V22.N2.47-56","url":null,"abstract":" Lactobacillus casei WSP-derived an aspartic protease was sequentially purified by using chromatography gel filtration sephadex G-50. It resulted in a 22.81-fold increase of specific activity (51.5 U/mg) with a final yield of 1.9%. The estimated molecular weight of the purified enzyme was 37 kDa and showed gelatinolytic activity in zymogram assay. The enzyme exhibited optimum activity at 40ºC and pH 6 with casein as the substrate. Enzyme activity was significantly inhibited by pepstatin A (0.5 mM and 1 mM), confirming that this enzyme is a group of aspartic proteases, while other inhibitors such as EDTA, PMSF and iodoacetic acid showed no inhibition effect on the activity of enzyme. The addition of metal ion to the enzyme decreased enzyme activity, indicating the proteolytic enzyme was metal ion- dependent. Denaturant such as DDT tended to increase caseinolytic activity. Furthermore, this enzyme was capable of generating the new peptides from skimmed milk with the size 8 kDa, 10 kDa and 15 kDa. These peptides have potential as antibacterial and antioxidant agents.","PeriodicalId":41037,"journal":{"name":"Annales Bogorienses-Journal of Tropical General Botany","volume":"14 10 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2018-12-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"82570607","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2018-12-31DOI: 10.14203/ANN.BOGOR.2018.V22.N2.65-74
Tarwadi Tarwadi, H. Rachmawati, R. E. Kartasasmita, S. Pambudi, A. D. Arbianto, D. Restiani, Sukmadjaja Asyarie
The main requirement of transfection agent has to condense DNA in nanoparticle size, protect the DNA from nucleases and other degrading enzymes during its transport in cell cytoplasm and nucleus and should not toxic to target cells. In this research, lipopeptide composed of palmitoyl (C-16) and short peptide sequence have been designed fully synthesized and tested to DNA condensation capability and toxicity. The DNA condensation study was performed using EtBr exclusion assay and cytotoxicity determination was carried out by colorimetric MTT assay. It was revealed that lipopeptide-based transfection agent of Pal-CKKHH and Pal-CKKHH-YGRKKRRQRRR-PKKKRKV condensed DNA molecules efficiently. The lipopeptide was less toxic compared to Lipofectamine and Poly-L-Lysine, that shown by 90% of CHO-K1 cells remained viable when they were treated with 4.36 µM Pal-CKKHHYGRKKRRQRRR-PKKKRKV. Meanwhile, there were only ~75% and 80% of CHO-K1 viable cells when it was treated with PLL and Lipofectamine®2000, respectively. Moreover, cell viability of HepG2 was ~ 75% after treated with 2.18 µM of Pal-CKKHH-YGRKKRRQRRR-PKKKRKV and decreased to ~65% when the lipopeptide concentration increased to 8.72 M. In summary, the synthesized lipopeptide condenses DNA molecules efficiently, less toxic than Lipofectamine®2000 and PLL and has possibility to be explored as a non-viral gene delivery vehicle.
{"title":"DNA Condensation Study of Fully Synthesized Lipopeptide-Based Transfection Agent for Gene Delivery Vehicle","authors":"Tarwadi Tarwadi, H. Rachmawati, R. E. Kartasasmita, S. Pambudi, A. D. Arbianto, D. Restiani, Sukmadjaja Asyarie","doi":"10.14203/ANN.BOGOR.2018.V22.N2.65-74","DOIUrl":"https://doi.org/10.14203/ANN.BOGOR.2018.V22.N2.65-74","url":null,"abstract":" The main requirement of transfection agent has to condense DNA in nanoparticle size, protect the DNA from nucleases and other degrading enzymes during its transport in cell cytoplasm and nucleus and should not toxic to target cells. In this research, lipopeptide composed of palmitoyl (C-16) and short peptide sequence have been designed fully synthesized and tested to DNA condensation capability and toxicity. The DNA condensation study was performed using EtBr exclusion assay and cytotoxicity determination was carried out by colorimetric MTT assay. It was revealed that lipopeptide-based transfection agent of Pal-CKKHH and Pal-CKKHH-YGRKKRRQRRR-PKKKRKV condensed DNA molecules efficiently. The lipopeptide was less toxic compared to Lipofectamine and Poly-L-Lysine, that shown by 90% of CHO-K1 cells remained viable when they were treated with 4.36 µM Pal-CKKHHYGRKKRRQRRR-PKKKRKV. Meanwhile, there were only ~75% and 80% of CHO-K1 viable cells when it was treated with PLL and Lipofectamine®2000, respectively. Moreover, cell viability of HepG2 was ~ 75% after treated with 2.18 µM of Pal-CKKHH-YGRKKRRQRRR-PKKKRKV and decreased to ~65% when the lipopeptide concentration increased to 8.72 M. In summary, the synthesized lipopeptide condenses DNA molecules efficiently, less toxic than Lipofectamine®2000 and PLL and has possibility to be explored as a non-viral gene delivery vehicle.","PeriodicalId":41037,"journal":{"name":"Annales Bogorienses-Journal of Tropical General Botany","volume":"1 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2018-12-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"82504584","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2018-12-31DOI: 10.14203/ann.bogor.2018.v22.n2.94-100
F. Fatimah, J. Prasetiyono, K. Trijatmiko, Sustiprijatno Sustiprijatno
The sustainability and increasing the national rice production require the readiness of food and agriculture sector cope with the impacts of climate change, land degradation, drought area, sloping production and the raising of population growth. Adaptation plays an important role in ensuring the sustainability of food security. This research aimed to develop drought-tolerant variety of Inpari 30 (submergence tolerance variety) and Situ Bagendit through marker-assisted backcrossing-through pyramiding gene of identified QTLs for foreground selection and to explore SSRs and 6K SNPs for background selection distributed in 12 rice chromosome of drought tolerant donor (Cabacu) and recipient rice (Inpari 30 and Situ Bagendit). The foreground selection revealed that flanking SSRs of each QTLs (qRPF2.1, qGPP2.1, qSPP4.1 and Sub1) was less than 2 cM. The background selection through polymorphic survey of Rice 6K SNP primers revealed 2457 (53,3%) polymorphic SNPs on Inpari 30 vs Cabacu and 2563 (55,6%) polymorphic SNPs on Situ Bagendit vs Cabacu with the average distance about 0.74 cM/chromosome. The genotypic selection of F1 Inpari 30/Cabacu and F1 Situ Bagendit/Cabacu have already in heterozygote condition for these 4 QTLs target. These lines was continued for backcross breeding to develop BC1F1 Inpari 30/Cabacu and BC1F1 Situ Bagendit/Cabacu generation.
{"title":"Molecular Evaluation for Drought Tolerant Using Marker Assisted Breeding Method","authors":"F. Fatimah, J. Prasetiyono, K. Trijatmiko, Sustiprijatno Sustiprijatno","doi":"10.14203/ann.bogor.2018.v22.n2.94-100","DOIUrl":"https://doi.org/10.14203/ann.bogor.2018.v22.n2.94-100","url":null,"abstract":" The sustainability and increasing the national rice production require the readiness of food and agriculture sector cope with the impacts of climate change, land degradation, drought area, sloping production and the raising of population growth. Adaptation plays an important role in ensuring the sustainability of food security. This research aimed to develop drought-tolerant variety of Inpari 30 (submergence tolerance variety) and Situ Bagendit through marker-assisted backcrossing-through pyramiding gene of identified QTLs for foreground selection and to explore SSRs and 6K SNPs for background selection distributed in 12 rice chromosome of drought tolerant donor (Cabacu) and recipient rice (Inpari 30 and Situ Bagendit). The foreground selection revealed that flanking SSRs of each QTLs (qRPF2.1, qGPP2.1, qSPP4.1 and Sub1) was less than 2 cM. The background selection through polymorphic survey of Rice 6K SNP primers revealed 2457 (53,3%) polymorphic SNPs on Inpari 30 vs Cabacu and 2563 (55,6%) polymorphic SNPs on Situ Bagendit vs Cabacu with the average distance about 0.74 cM/chromosome. The genotypic selection of F1 Inpari 30/Cabacu and F1 Situ Bagendit/Cabacu have already in heterozygote condition for these 4 QTLs target. These lines was continued for backcross breeding to develop BC1F1 Inpari 30/Cabacu and BC1F1 Situ Bagendit/Cabacu generation.","PeriodicalId":41037,"journal":{"name":"Annales Bogorienses-Journal of Tropical General Botany","volume":"38 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2018-12-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"82190745","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2018-06-29DOI: 10.14203/ANN.BOGOR.2018.V22.N1.12-19
A. Thontowi, E. Yetti, Yopi Yopi
Alkanes are major component of crude oil that could be hydrolyzed by the enzyme of alkane hydroxylase. The are three types of alkane hydroxylase based on the chain length of alkane such as short-chain length/SCL (C2-C4), medium-chain length/MCL (C5-C17), and long-chain length/LCL (C>18). The aims of this study were to characterize and identify alkanes-degrading bacteria from these bacteria. The 30 strains from marine were grown on MCL (Pentane-C5H12, Decane-C10H22, and Pentadecane-C15H32) and LCL (n-Paraffin-C12H19C17 and branch of Pristane-C19H40). The study showed twenty-nine isolates have the ability to degrade alkanes compounds, whereas 14 isolates have grown ability on MCL and LCL medium, 11 isolates have the ability to grow on MCL and n-LCL, 3 isolates have the ability only to grow on MCL medium and 1 isolate has the ability only grow on n-LCL medium. The growth test result indicated that 29 isolates have medium-chain alkane monooxygenase and long-chain alkane hydroxylase. Based on 16S rDNA gene analysis, we obtained twenty nine of oil- degrading bacteria, namely a-proteobacteria (57 %), g-proteobacteria (30 %), Flavobacteria (7 %), Bacilli (3%) and Propionibacteriales (3 %). g-Proteobacteria and a-proteobacteria which seems to play an important role in the alkane biodegradation.
{"title":"Medium Chain and Long Chain Alkanes Hydroxylase Producing Whole Cell Biocatalyst From Marine Bacteria","authors":"A. Thontowi, E. Yetti, Yopi Yopi","doi":"10.14203/ANN.BOGOR.2018.V22.N1.12-19","DOIUrl":"https://doi.org/10.14203/ANN.BOGOR.2018.V22.N1.12-19","url":null,"abstract":"Alkanes are major component of crude oil that could be hydrolyzed by the enzyme of alkane hydroxylase. The are three types of alkane hydroxylase based on the chain length of alkane such as short-chain length/SCL (C2-C4), medium-chain length/MCL (C5-C17), and long-chain length/LCL (C>18). The aims of this study were to characterize and identify alkanes-degrading bacteria from these bacteria. The 30 strains from marine were grown on MCL (Pentane-C5H12, Decane-C10H22, and Pentadecane-C15H32) and LCL (n-Paraffin-C12H19C17 and branch of Pristane-C19H40). The study showed twenty-nine isolates have the ability to degrade alkanes compounds, whereas 14 isolates have grown ability on MCL and LCL medium, 11 isolates have the ability to grow on MCL and n-LCL, 3 isolates have the ability only to grow on MCL medium and 1 isolate has the ability only grow on n-LCL medium. The growth test result indicated that 29 isolates have medium-chain alkane monooxygenase and long-chain alkane hydroxylase. Based on 16S rDNA gene analysis, we obtained twenty nine of oil- degrading bacteria, namely a-proteobacteria (57 %), g-proteobacteria (30 %), Flavobacteria (7 %), Bacilli (3%) and Propionibacteriales (3 %). g-Proteobacteria and a-proteobacteria which seems to play an important role in the alkane biodegradation.","PeriodicalId":41037,"journal":{"name":"Annales Bogorienses-Journal of Tropical General Botany","volume":"55 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2018-06-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"86259526","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2018-06-29DOI: 10.14203/ANN.BOGOR.2018.V22.N1.27-34
F. Fatimah, J. Prasetiyono, A. Polosoro, M. Baroya
Rice bacterial leaf blight (BLB) disease caused by Xanthomonas oryzae pv. oryzae (Xoo) affected grain yield and decreasing rice production in rice growing countries. Conde, Indonesian rice variety, exhibits high resistance to most of the Indonesian races of (BLB) and has been used in Indonesia for cultivated rice. This study was aimed to conduct the molecular detection between proximal markers in chromosome 6 and relative expression of Conde rice variety compare to IRBB7 in Xa7 region. The population screening, BLB evaluation and molecular detection around the Xa7 region were conducted. The results showed from the collection of individual recombinants between resistant and susceptible parents narrow the region containing the BTBPOZ domain. The sequence alignment of Xa7LD37 in two resistant and three susceptible cultivars demonstrated a perfect association. The sequence alignment in exon region of Loc_Os06g46240 in Nipponbare, IRBB7, and IR64 identified indel/SNPs in this region leading to nucleotide substitution and frameshift resulting in amino acid change between resistant and susceptible cultivars. It was predicted that Conde revealed the similar gene action with Xa7 gene for BLB that encodes a BTB POZ domain.
{"title":"Molecular Detection of Resistance To Bacterial Leaf Blight on Conde Indonesian Rice Variety","authors":"F. Fatimah, J. Prasetiyono, A. Polosoro, M. Baroya","doi":"10.14203/ANN.BOGOR.2018.V22.N1.27-34","DOIUrl":"https://doi.org/10.14203/ANN.BOGOR.2018.V22.N1.27-34","url":null,"abstract":"Rice bacterial leaf blight (BLB) disease caused by Xanthomonas oryzae pv. oryzae (Xoo) affected grain yield and decreasing rice production in rice growing countries. Conde, Indonesian rice variety, exhibits high resistance to most of the Indonesian races of (BLB) and has been used in Indonesia for cultivated rice. This study was aimed to conduct the molecular detection between proximal markers in chromosome 6 and relative expression of Conde rice variety compare to IRBB7 in Xa7 region. The population screening, BLB evaluation and molecular detection around the Xa7 region were conducted. The results showed from the collection of individual recombinants between resistant and susceptible parents narrow the region containing the BTBPOZ domain. The sequence alignment of Xa7LD37 in two resistant and three susceptible cultivars demonstrated a perfect association. The sequence alignment in exon region of Loc_Os06g46240 in Nipponbare, IRBB7, and IR64 identified indel/SNPs in this region leading to nucleotide substitution and frameshift resulting in amino acid change between resistant and susceptible cultivars. It was predicted that Conde revealed the similar gene action with Xa7 gene for BLB that encodes a BTB POZ domain.","PeriodicalId":41037,"journal":{"name":"Annales Bogorienses-Journal of Tropical General Botany","volume":"46 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2018-06-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"73841420","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2018-06-29DOI: 10.14203/ANN.BOGOR.2018.V22.N1.35-46
Supatmi Supatmi, N. Rahman, N. Hartati
Cassava (Manihot esculenta) is an important crop to food security under climate change due to its various tolerance mechanism under stress conditions. However, the sustainable growth of cassava in the field depends on many factors especially temperature. The objective of the research was to investigate the growth performances and proline contents of irradiated Ubi Kuning at dosage of 10 Gy, cultured in Murashige Skoog (MS) hormone-free solid medium for 4 weeks at three different temperature treatments i.e 25oC, 28oC and 30oC. Each treatment consisted of 3 clone explants with 5 replicates. Results show that growth performances of irradiated plantlets were better compared to that of non-irradiated plantlets in terms of plant height and number of leaves at all temperature tested. The best growth performances were obtained from irradiated plantlets grown under 30oC. The proline content of irradiated Ubi Kuning was high when they were grown under 25 oC and 30oC, implying that these plantlets had the possibility to tolerant to lower and higher-temperature condition. This study is initially useful to find out the growth ability of irradiated Ubi Kuning in response to lower and higher temperature.
{"title":"Growth and Proline Content of Irradiated in Vitro Shoots of Ubi Kuning Cassava Genotype Cultured at Different Temperatures","authors":"Supatmi Supatmi, N. Rahman, N. Hartati","doi":"10.14203/ANN.BOGOR.2018.V22.N1.35-46","DOIUrl":"https://doi.org/10.14203/ANN.BOGOR.2018.V22.N1.35-46","url":null,"abstract":"Cassava (Manihot esculenta) is an important crop to food security under climate change due to its various tolerance mechanism under stress conditions. However, the sustainable growth of cassava in the field depends on many factors especially temperature. The objective of the research was to investigate the growth performances and proline contents of irradiated Ubi Kuning at dosage of 10 Gy, cultured in Murashige Skoog (MS) hormone-free solid medium for 4 weeks at three different temperature treatments i.e 25oC, 28oC and 30oC. Each treatment consisted of 3 clone explants with 5 replicates. Results show that growth performances of irradiated plantlets were better compared to that of non-irradiated plantlets in terms of plant height and number of leaves at all temperature tested. The best growth performances were obtained from irradiated plantlets grown under 30oC. The proline content of irradiated Ubi Kuning was high when they were grown under 25 oC and 30oC, implying that these plantlets had the possibility to tolerant to lower and higher-temperature condition. This study is initially useful to find out the growth ability of irradiated Ubi Kuning in response to lower and higher temperature.","PeriodicalId":41037,"journal":{"name":"Annales Bogorienses-Journal of Tropical General Botany","volume":"81 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2018-06-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"75247542","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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