Benzylsuccinate Synthase is Post-Transcriptionally Regulated in the Toluene-Degrading Denitrifier Magnetospirillum sp. Strain 15-1
Abstract
:1. Introduction
2. Materials and Methods
2.1. Growth Medium and Cultivation
2.2. RNA Extraction and cDNA Synthesis
2.3. Transcriptional Organization of bss, tdi, and bbs Gene Clusters by Reverse-Transcriptase PCR (RT-PCR)
2.4. Gene Expression Assessment by Quantitative RT- PCR (qRT-PCR)
2.5. Protein Extraction and Preparation
2.6. LC-MS/MS and Proteome Analysis
2.7. Data Availability
3. Results
3.1. Genomic Organization of Genes Involved in Anaerobic Toluene Transformation
3.2. Regulation of bss Gene Expression in Magnetospirillum sp. Strain 15-1
3.3. Proteins Involved in Anaerobic Toluene Degradation
3.4. Expression Pattern of the Transcriptional Regulatory Components TdiCRS and XylR
3.5. Proteins Involved in the Denitrification Process
4. Discussion
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Primers Used for PCR | ||||
Target Sequence | Primer Name | Primer Sequence | Product Size (bp) | Source |
16S rRNA | 27F 1492R | AGAGTTTGATCMTGGCTCAG GGYTACCTTGTTACGACTT | 1465 | [31] |
bssA | bssA_F1 bssA_R2 | GACGARTTCATCRTCGGCTACCACGC AGCAGRTTGSCYTTCTGRTTYTTCTG | 1546 | Junca, H. |
bcrC | bcrC_F bcrC_R | CGHATYCCRCGSTCGACCATCG CGGATCGGCTGCATCTGGCC | 800 | [32] |
bbsC region | bbsD_F bbsB_R | GGCGGGATGTTGTCCTATGG GCTTCGGCCCTATTTGCTTG | 1823 | this study |
Primers Used for RT-qPCR | ||||
Target Sequence | Primer Name | Primer Sequence | Product Size (bp) | Source |
16S rRNA | 16S_F 16S_R | TGATGAAGGCCTTAGGGTTG CCAGGGCTTTCACTTCTGAC | 170 | Marín, V. |
bcrC | 2bcrC_F 2bcrC_R | CATGATCTTCCCGTT TCCTTCAGCTCCTTC | 159 | Marín, V. |
bssA | bssA_F3 bssA_R3 | CGTCCTTCGCCTCGGGTTAC CATCGCCTGCCAGTTGTCAATC | 188 | Marín, V. |
Primers Used for RT-PCR | ||||
Target Sequence | Primer Name | Primer Sequence | Product Size (bp) | Source |
Intergenic region bssD-bssA | bssD-bssA_F bssD-bssA_R | CGCATTCACATCCCGGTCATC CAGGACGTTGGCGGTCATATT | 575 | this study |
Intergenic region bssA-bssE | bssA-bssE_F bssA-bssE_R | CTGAATTGCGACCTCTGAGC GCTTGCGCGAGATATTACGG | 587 | this study |
Intergenic region bssE-bssF | bssE-bssF_F bssE-bssF_R | GGCAGGTCTGGATGGATGAG CGTTTGGGAGGGGTATCGTC | 504 | this study |
Intergenic region bssF-bssG | bssF-bssG_F bssF-bssG_R | GCCATTCGCCGTTTCAACAA GCGAACCTGGGAAAACATCG | 482 | this study |
Intergenic region bssG-bssP | bssG-bssP_F bssG-bssP_R | CCTTGGATGAAGGCCGTAAC CCGAAGATCAGCCACTTTCC | 560 | this study |
Intergenic region bssP-bssI | bssP-bssI_F bssP-bssI_R | CCCGTTTCAAGCGATGGTTC GAACTCCCTCCTTGCGGTAG | 493 | this study |
Intergenic region bssI-bssJ | bssI-bssJ_F1 bssI-bssJ_R1 | CGAATTCCCCAGCGACTCA ATCTCGAACACGCCCACTC | 561 | this study |
Intergenic region bssJ-bssK | bssJ-bssK_F bssJ-bssK_R | GCCCCCTACTTCAAATGGCT GTATCCCAATCCAGCGGAGG | 480 | this study |
Intergenic region bssK-bssL | bssK-bssL_F bssK-bssL_R | AGTTGATTTCCTACGGGCGG GTGAGCAATCCACATGACGC | 515 | this study |
Intergenic region tdiC-tdiR | tdiC-tdiR_F tdiC-tdiR_R | GAGATCATGACGACGGAGGT TAGAAAGATCAGCGGCAGCG | 500 | this study |
Intergenic region tdiR-tdiS | tdiR-tdiS_F tdiR-tdiS_R | CGCCTCAGCAAGGAAGTGTT GGAAGCAAATGCCAACGGG | 435 | this study |
xylR | xylR_F xylR_R | TGTCGAGCGTGGCTATTACTC CTTCCACCAAATTCTCGGGC | 545 | this study |
Intergenic region bbsA-bbsB | bbsA-bbsB_F bbsA-bbsB_R | GGGCAGCTTGATTTTCCCAA GGCGATGAACACATCTCGTTG | 436 | this study |
Intergenic region bbsB-bbsC | bbsB-bbsC_F bbsB-bbsC_R | GGCGGGATGTTGTCCTATGG ACCGATTCCGGAAGGAAAGG | 535 | this study |
Intergenic region bbsC-bbsD | bbsC-bbsD_F bbsC-bbsD_R | CGGCAAGAGCGCCTATTTCT GGTGATTCCATTGCGTCCCA | 611 | this study |
Intergenic region bbsD-bbsE | bbsD-bbsE_F bbsD-bbsE_R | CTGGGACGCAATGGAATCAC GCGCAAAAACACCTCCCTG | 542 | this study |
Intergenic region bbsE-bbsF | bbsE-bbsF_F1 bbsE-bbsF_R1 | ATACGCCTATCGGACCTCGG GCTCTGCATGAACCACTTCC | 380 | this study |
Intergenic region bbsF-bbsG | bbsF-bbsG_F bbsF-bbsG_R | CGCGGTGTTTTCCGATGAAG TTCAGCATTGCGTGCTCTTG | 550 | this study |
Intergenic region bbsG-bbsH | bbsG-bbsH_F bbsG-bbsH_R | CGGGAAACCGGCATCGAATA TTCATCTGGGGGATGAGGGG | 428 | this study |
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Meyer-Cifuentes, I.; Gruhl, S.; Haange, S.-B.; Lünsmann, V.; Jehmlich, N.; von Bergen, M.; Heipieper, H.J.; Müller, J.A. Benzylsuccinate Synthase is Post-Transcriptionally Regulated in the Toluene-Degrading Denitrifier Magnetospirillum sp. Strain 15-1. Microorganisms 2020, 8, 681. https://doi.org/10.3390/microorganisms8050681
Meyer-Cifuentes I, Gruhl S, Haange S-B, Lünsmann V, Jehmlich N, von Bergen M, Heipieper HJ, Müller JA. Benzylsuccinate Synthase is Post-Transcriptionally Regulated in the Toluene-Degrading Denitrifier Magnetospirillum sp. Strain 15-1. Microorganisms. 2020; 8(5):681. https://doi.org/10.3390/microorganisms8050681
Chicago/Turabian StyleMeyer-Cifuentes, Ingrid, Sylvie Gruhl, Sven-Bastiaan Haange, Vanessa Lünsmann, Nico Jehmlich, Martin von Bergen, Hermann J. Heipieper, and Jochen A. Müller. 2020. "Benzylsuccinate Synthase is Post-Transcriptionally Regulated in the Toluene-Degrading Denitrifier Magnetospirillum sp. Strain 15-1" Microorganisms 8, no. 5: 681. https://doi.org/10.3390/microorganisms8050681