Apiradee Hongsthong - Academia.edu (original) (raw)
Papers by Apiradee Hongsthong
Nondenaturing polyacrylamide gel electrophoresis and activity staining 66 Determination of molecu... more Nondenaturing polyacrylamide gel electrophoresis and activity staining 66 Determination of molecular weight of ATCase 69 Sodium dodecyl sulfate denaturing polyacrylamide gel electrophoresis 71 Electrocution 74 Hummel-Dreyer method 76 Densitometry 79 RESULTS 88 Carbamoylaspartate standard curve 88 Protein standard curve 88 Transformants containing plasmids 91 Partial purification of the ATCase from transformants 91 Kinetic and effector response of the ATCase hybrids 95 Analysis of the ATCase hybrids 97 Protein-protein binding studies by Hummel-Dreyer method.. .. 114 Analysis of the P.putida WT ATCase after denaturation and renaturation 115 Determination of the protein ratios in the assembly of the P.putida
CRC Press eBooks, Jul 12, 2019
Algae and their Biotechnological Potential, 2001
The cyanobacterium Spirulina platensis strain C 1 contains three desaturases, ∆9,∆12 and ∆6. Thes... more The cyanobacterium Spirulina platensis strain C 1 contains three desaturases, ∆9,∆12 and ∆6. These desaturases introduce double bonds at the ∆9, ∆12 and ∆6 position of fatty acids that have been esterified to glycerolipids. In cyanobacteria, the elevation of unsaturated fatty acid level in membrane lipids has been shown to play a major role in response to temperature change. We therefore carried out a study of the regulation of desaturase gene expression in response to temperature change in S. platensis strain C 1. In a previous study, it was found that the transcription of desC (∆9) and desA (∆ 12) was not affected by a temperature shift from 35 °C (normal growth temperature) to 22 °C or to 40 °C. However, while desC and desA remained unaffected, the transcription and mRNA stability of desD (∆6) were enhanced by lowering the temperature from 35 °C to 22 °C. In this study, these genes were studied at the translation level by Western blot analysis. Two synthetic peptides of ∆9 and ∆6 desaturases w1th 20 amino acids in length from the Cterminus were synthesized. Polyclonal antibodies, which were raised against these synthetic peptides, were used in the Western blot analysis. The level of ∆9 desaturase was not significantly affected in either thylakoid or plasma membrane after the temperature shift. By contrast, it was found that ∆6 desaturase increased in thylakoid membrane after lowering the temperature from 35 °C to 22 °C for 24 h, although it remained constant in plasma membrane. These results indicate that the expression of the ∆6desaturase gene is up regulated by temperature in different ways in two types of lipid membranes, while that of ∆9desatura.se is temperature independent.
Abstract Algae and cyanobacteria have been studied for their feasibility to use as a source for p... more Abstract Algae and cyanobacteria have been studied for their feasibility to use as a source for production of bioproduct(s) due to their ability to absorb greenhouse gas (carbon dioxide) and to be grown large-scale outdoor cultures. Spirulina is the only cyanobacterium that can be mass cultivated in outdoor ponds, thus it can be a model organism for biofactory. Genome-scale study is relevant for screening of bioproduct synthetic capability, whereas proteome-wide analyses for protein expression level and posttranslational modification lead to information required for understanding of cellular response and regulation under abiotic stress conditions that the cells encounter in outdoor mass cultivation system. Here, we present our current understanding of protein networks in Spirulina, their regulation under temperature stress, and bioinformatics tools that are used for characterizing them. We then discuss the involvement of molecular chaperones, which play a vital role in protein quality control under stress conditions, in cellular protein networks.
BMC Molecular and Cell Biology
Synechocystis histidine kinase, Sll0474: Hik28, a signal protein in a two-component signal transd... more Synechocystis histidine kinase, Sll0474: Hik28, a signal protein in a two-component signal transduction system, plays a critical role in responding to a decrease in growth temperature and is also involved in nitrogen metabolism. In the present study, under combined stress from non-optimal growth temperature and nitrogen depletion, a comparative proteomic analysis of the wild type (WT) and a deletion mutant (MT) of Synechocystis histidine kinase, Sll0474: Hik28, in a two-component signal transduction system identified the specific groups of ABC transporters that were Hik28-dependent, e.g., the iron transporter, and Hik28-independent, e.g., the phosphate transporter. The iron transporter, AfuA, was found to be upregulated only in the WT strain grown under the combined stress of high temperature and nitrogen depletion. Whereas, the expression level of the phosphate transporter, PstS, was increased in both the WT and MT strains. Moreover, the location in the genome of the genes encoding...
Frontiers in Genetics
Hypertension or elevated blood pressure is a serious medical condition that significantly increas... more Hypertension or elevated blood pressure is a serious medical condition that significantly increases the risks of cardiovascular disease, heart disease, diabetes, stroke, kidney disease, and other health problems, that affect people worldwide. Thus, hypertension is one of the major global causes of premature death. Regarding the prevention and treatment of hypertension with no or few side effects, antihypertensive peptides (AHTPs) obtained from natural sources might be useful as nutraceuticals. Therefore, the search for alternative/novel AHTPs in food or natural sources has received much attention, as AHTPs may be functional agents for human health. AHTPs have been observed in diverse organisms, although many of them remain underinvestigated. The identification of peptides with antihypertensive activity in the laboratory is time- and resource-consuming. Alternatively, computational methods based on robust machine learning can identify or screen potential AHTP candidates prior to expe...
Additional file 9. Protein-protein interaction analyses of the proteins of interest by using yeas... more Additional file 9. Protein-protein interaction analyses of the proteins of interest by using yeast two-hybrid system (Y2H).
Additional file 4. MS/MS spectra of some peptides of interest.
Additional file 2. PPI subnetworks of four groups of the differentially expressed proteins; (A) p... more Additional file 2. PPI subnetworks of four groups of the differentially expressed proteins; (A) proteins upregulated at 22 °C and downregulated at 40 °C, (B) proteins downregulated at 22 °C and upregulated at 40 °C, (C) proteins upregulated at both temperatures (D) proteins downregulated at both temperatures. The subnetworks were constructed by using STRING. The A. platensis C1 proteins were inferred to that of the A. platensis NIES39 via orthologous group.
Additional file 16. List of the 527 phosphorylated proteins after filtering by the high stringenc... more Additional file 16. List of the 527 phosphorylated proteins after filtering by the high stringency criteria.
Additional file 19. Pictures of pGBKT7 and pGADT7 AD vectors (Clontech Laboratories, Inc. USA).
Additional file 3. List of 101 proteins after the threshold filtering.
Additional file 15. PPI subnetworks of the proteins in the same orthologous group as (A) SPLC1_S0... more Additional file 15. PPI subnetworks of the proteins in the same orthologous group as (A) SPLC1_S082010, (B) SPLC1_S540750 and (C) SPLC1_S360070. The subnetworks were constructed by using STRING. The A. platensis C1 proteins were inferred to that of the A. platensis NIES39 via orthologous group.
Additional file 10. Construction of Δsll0474 mutant in Synechocystis sp. PCC6803; (A) Map of the ... more Additional file 10. Construction of Δsll0474 mutant in Synechocystis sp. PCC6803; (A) Map of the recombinant plasmid containing the upstream (us) and the downstream (ds) regions of the sll0474 gene, and the spectinomycin resistant gene (Spec) cloned in pGEM T-Easy vector (B) Enzymatic digestion of the recombinant plasmids as shown in (A) represented the inserts (The recombinant plasmids in the last two lanes of the agarose gel were digested with EcoRI&HindIII and PstI, respectively.) (C) PCR products of Synechocystis sp. 6803WT and Δsll0474 mutant amplified by using US-sll0474ERIFW and DS-sll0474pstIRVprimers (D) PCR products of Synechocystis sp. 6803WT and Δsll0474 mutant amplified by using Sll0474ERIFW and Sll0474BaHIRV primers.
Table S1. Dataset and output comparison between SpirPep and BIOPEP, the identified bioactive pept... more Table S1. Dataset and output comparison between SpirPep and BIOPEP, the identified bioactive peptides obtained from temperature stress â expressed protein [30] digested with trypsin and no miscleavage. (XLSX 18Â kb)
Figure S3. Example of webpage snapshots from the SpirPep tool: a) query input from user submissio... more Figure S3. Example of webpage snapshots from the SpirPep tool: a) query input from user submission (protein sequences, userâ s name and email, job title, desired enzyme(s) and allowed miscleavage number), b) notification of received query from users as the response page and submission information email when the analysis complete, and c) results pages that contain the list all and summary pages, which show the predicted bioactive peptide sequences derived from input proteins and parameters. This provides a valuable decision to users for their re-designing the digestion system to obtain desirable bioactive peptides and enzymes before validation in a laboratory. (JPEG 2333Â kb)
Arthrospira platensis is a cyanobacterium that is extensively cultivated outdoors on a large comm... more Arthrospira platensis is a cyanobacterium that is extensively cultivated outdoors on a large commercial scale for consumption as a food for humans and animals. It can be grown in monoculture under highly alkaline conditions, making it attractive for industrial production. Here we describe the complete genome sequence of A. platensis C1 strain and its annotation. The A. platensis C1 genome contains 6,089,210 bp including 6,108 protein-coding genes and 45 RNA genes, and no plasmids. The genome information has been used for further comparative analysis, particularly of metabolic pathways, photosynthetic efficiency and barriers to gene transfer.
Comparative analysis of the Spirulina platensis subcellular proteome in response to low- and high... more Comparative analysis of the Spirulina platensis subcellular proteome in response to low- and high-temperature stresses: uncovering cross-talk of signaling components
Additional file 7. PPI subnetwork of Hik28 and enzymes involved in fatty acid desaturation. The s... more Additional file 7. PPI subnetwork of Hik28 and enzymes involved in fatty acid desaturation. The subnetwork was constructed by using STRING. The A. platensis C1 proteins were inferred to that of the Synechocystis sp. PCC6803 via orthologous group.
Nondenaturing polyacrylamide gel electrophoresis and activity staining 66 Determination of molecu... more Nondenaturing polyacrylamide gel electrophoresis and activity staining 66 Determination of molecular weight of ATCase 69 Sodium dodecyl sulfate denaturing polyacrylamide gel electrophoresis 71 Electrocution 74 Hummel-Dreyer method 76 Densitometry 79 RESULTS 88 Carbamoylaspartate standard curve 88 Protein standard curve 88 Transformants containing plasmids 91 Partial purification of the ATCase from transformants 91 Kinetic and effector response of the ATCase hybrids 95 Analysis of the ATCase hybrids 97 Protein-protein binding studies by Hummel-Dreyer method.. .. 114 Analysis of the P.putida WT ATCase after denaturation and renaturation 115 Determination of the protein ratios in the assembly of the P.putida
CRC Press eBooks, Jul 12, 2019
Algae and their Biotechnological Potential, 2001
The cyanobacterium Spirulina platensis strain C 1 contains three desaturases, ∆9,∆12 and ∆6. Thes... more The cyanobacterium Spirulina platensis strain C 1 contains three desaturases, ∆9,∆12 and ∆6. These desaturases introduce double bonds at the ∆9, ∆12 and ∆6 position of fatty acids that have been esterified to glycerolipids. In cyanobacteria, the elevation of unsaturated fatty acid level in membrane lipids has been shown to play a major role in response to temperature change. We therefore carried out a study of the regulation of desaturase gene expression in response to temperature change in S. platensis strain C 1. In a previous study, it was found that the transcription of desC (∆9) and desA (∆ 12) was not affected by a temperature shift from 35 °C (normal growth temperature) to 22 °C or to 40 °C. However, while desC and desA remained unaffected, the transcription and mRNA stability of desD (∆6) were enhanced by lowering the temperature from 35 °C to 22 °C. In this study, these genes were studied at the translation level by Western blot analysis. Two synthetic peptides of ∆9 and ∆6 desaturases w1th 20 amino acids in length from the Cterminus were synthesized. Polyclonal antibodies, which were raised against these synthetic peptides, were used in the Western blot analysis. The level of ∆9 desaturase was not significantly affected in either thylakoid or plasma membrane after the temperature shift. By contrast, it was found that ∆6 desaturase increased in thylakoid membrane after lowering the temperature from 35 °C to 22 °C for 24 h, although it remained constant in plasma membrane. These results indicate that the expression of the ∆6desaturase gene is up regulated by temperature in different ways in two types of lipid membranes, while that of ∆9desatura.se is temperature independent.
Abstract Algae and cyanobacteria have been studied for their feasibility to use as a source for p... more Abstract Algae and cyanobacteria have been studied for their feasibility to use as a source for production of bioproduct(s) due to their ability to absorb greenhouse gas (carbon dioxide) and to be grown large-scale outdoor cultures. Spirulina is the only cyanobacterium that can be mass cultivated in outdoor ponds, thus it can be a model organism for biofactory. Genome-scale study is relevant for screening of bioproduct synthetic capability, whereas proteome-wide analyses for protein expression level and posttranslational modification lead to information required for understanding of cellular response and regulation under abiotic stress conditions that the cells encounter in outdoor mass cultivation system. Here, we present our current understanding of protein networks in Spirulina, their regulation under temperature stress, and bioinformatics tools that are used for characterizing them. We then discuss the involvement of molecular chaperones, which play a vital role in protein quality control under stress conditions, in cellular protein networks.
BMC Molecular and Cell Biology
Synechocystis histidine kinase, Sll0474: Hik28, a signal protein in a two-component signal transd... more Synechocystis histidine kinase, Sll0474: Hik28, a signal protein in a two-component signal transduction system, plays a critical role in responding to a decrease in growth temperature and is also involved in nitrogen metabolism. In the present study, under combined stress from non-optimal growth temperature and nitrogen depletion, a comparative proteomic analysis of the wild type (WT) and a deletion mutant (MT) of Synechocystis histidine kinase, Sll0474: Hik28, in a two-component signal transduction system identified the specific groups of ABC transporters that were Hik28-dependent, e.g., the iron transporter, and Hik28-independent, e.g., the phosphate transporter. The iron transporter, AfuA, was found to be upregulated only in the WT strain grown under the combined stress of high temperature and nitrogen depletion. Whereas, the expression level of the phosphate transporter, PstS, was increased in both the WT and MT strains. Moreover, the location in the genome of the genes encoding...
Frontiers in Genetics
Hypertension or elevated blood pressure is a serious medical condition that significantly increas... more Hypertension or elevated blood pressure is a serious medical condition that significantly increases the risks of cardiovascular disease, heart disease, diabetes, stroke, kidney disease, and other health problems, that affect people worldwide. Thus, hypertension is one of the major global causes of premature death. Regarding the prevention and treatment of hypertension with no or few side effects, antihypertensive peptides (AHTPs) obtained from natural sources might be useful as nutraceuticals. Therefore, the search for alternative/novel AHTPs in food or natural sources has received much attention, as AHTPs may be functional agents for human health. AHTPs have been observed in diverse organisms, although many of them remain underinvestigated. The identification of peptides with antihypertensive activity in the laboratory is time- and resource-consuming. Alternatively, computational methods based on robust machine learning can identify or screen potential AHTP candidates prior to expe...
Additional file 9. Protein-protein interaction analyses of the proteins of interest by using yeas... more Additional file 9. Protein-protein interaction analyses of the proteins of interest by using yeast two-hybrid system (Y2H).
Additional file 4. MS/MS spectra of some peptides of interest.
Additional file 2. PPI subnetworks of four groups of the differentially expressed proteins; (A) p... more Additional file 2. PPI subnetworks of four groups of the differentially expressed proteins; (A) proteins upregulated at 22 °C and downregulated at 40 °C, (B) proteins downregulated at 22 °C and upregulated at 40 °C, (C) proteins upregulated at both temperatures (D) proteins downregulated at both temperatures. The subnetworks were constructed by using STRING. The A. platensis C1 proteins were inferred to that of the A. platensis NIES39 via orthologous group.
Additional file 16. List of the 527 phosphorylated proteins after filtering by the high stringenc... more Additional file 16. List of the 527 phosphorylated proteins after filtering by the high stringency criteria.
Additional file 19. Pictures of pGBKT7 and pGADT7 AD vectors (Clontech Laboratories, Inc. USA).
Additional file 3. List of 101 proteins after the threshold filtering.
Additional file 15. PPI subnetworks of the proteins in the same orthologous group as (A) SPLC1_S0... more Additional file 15. PPI subnetworks of the proteins in the same orthologous group as (A) SPLC1_S082010, (B) SPLC1_S540750 and (C) SPLC1_S360070. The subnetworks were constructed by using STRING. The A. platensis C1 proteins were inferred to that of the A. platensis NIES39 via orthologous group.
Additional file 10. Construction of Δsll0474 mutant in Synechocystis sp. PCC6803; (A) Map of the ... more Additional file 10. Construction of Δsll0474 mutant in Synechocystis sp. PCC6803; (A) Map of the recombinant plasmid containing the upstream (us) and the downstream (ds) regions of the sll0474 gene, and the spectinomycin resistant gene (Spec) cloned in pGEM T-Easy vector (B) Enzymatic digestion of the recombinant plasmids as shown in (A) represented the inserts (The recombinant plasmids in the last two lanes of the agarose gel were digested with EcoRI&HindIII and PstI, respectively.) (C) PCR products of Synechocystis sp. 6803WT and Δsll0474 mutant amplified by using US-sll0474ERIFW and DS-sll0474pstIRVprimers (D) PCR products of Synechocystis sp. 6803WT and Δsll0474 mutant amplified by using Sll0474ERIFW and Sll0474BaHIRV primers.
Table S1. Dataset and output comparison between SpirPep and BIOPEP, the identified bioactive pept... more Table S1. Dataset and output comparison between SpirPep and BIOPEP, the identified bioactive peptides obtained from temperature stress â expressed protein [30] digested with trypsin and no miscleavage. (XLSX 18Â kb)
Figure S3. Example of webpage snapshots from the SpirPep tool: a) query input from user submissio... more Figure S3. Example of webpage snapshots from the SpirPep tool: a) query input from user submission (protein sequences, userâ s name and email, job title, desired enzyme(s) and allowed miscleavage number), b) notification of received query from users as the response page and submission information email when the analysis complete, and c) results pages that contain the list all and summary pages, which show the predicted bioactive peptide sequences derived from input proteins and parameters. This provides a valuable decision to users for their re-designing the digestion system to obtain desirable bioactive peptides and enzymes before validation in a laboratory. (JPEG 2333Â kb)
Arthrospira platensis is a cyanobacterium that is extensively cultivated outdoors on a large comm... more Arthrospira platensis is a cyanobacterium that is extensively cultivated outdoors on a large commercial scale for consumption as a food for humans and animals. It can be grown in monoculture under highly alkaline conditions, making it attractive for industrial production. Here we describe the complete genome sequence of A. platensis C1 strain and its annotation. The A. platensis C1 genome contains 6,089,210 bp including 6,108 protein-coding genes and 45 RNA genes, and no plasmids. The genome information has been used for further comparative analysis, particularly of metabolic pathways, photosynthetic efficiency and barriers to gene transfer.
Comparative analysis of the Spirulina platensis subcellular proteome in response to low- and high... more Comparative analysis of the Spirulina platensis subcellular proteome in response to low- and high-temperature stresses: uncovering cross-talk of signaling components
Additional file 7. PPI subnetwork of Hik28 and enzymes involved in fatty acid desaturation. The s... more Additional file 7. PPI subnetwork of Hik28 and enzymes involved in fatty acid desaturation. The subnetwork was constructed by using STRING. The A. platensis C1 proteins were inferred to that of the Synechocystis sp. PCC6803 via orthologous group.