Current Research

Bacterial and plant enzymes for mimosine degradation. Mimosine [ß-N-(3-hydroxy-4-pyridone)-aminopropionic acid] is a toxin found in large quantities in the seeds, foliage and roots of Leucaena leucocephala (leucaena). Mimosine is toxic because it chelates several bivalent metallic ions and also because it forms a stable complex with pyridoxal-5’-phosphate (PLP). It inactivates a wide variety of enzymes by limiting their metallic ion cofactors and PLP coenzyme. Mimosine toxicity in animals causes various physiological abnormalities including enlarged thyroid glands, fetal deformities, and hair loss.  Rhizobium species strain TAL1145 that effectively nodulates leucaena can overcome mimosine toxicity by degrading it, and using the degradation products as a source of nutrients.  We are currently working on biochemistry of the enzymes encoded by the Rhizobium genes for mimosine degradation.  Interestingly, leucaena foliage also contains enzymes for mimosine degradation.  We are also characterizing the mimosine-degrading properties of one such lecuaena enzyme.

Genetic transformation of Leucaena leucocephala. As a woody tree-legume, leucaena is recalcitrant for genetic transformation. Recently, we have developed a protocol for genetic transformation of leucaena.  We are currently working on further improvement of the transformation protocol.  Previously, we transformed leucaena with the pydA gene from Rhizobium sp. strain TAL1145.  We are currently tranforming leucaena with a pydA-pydB fusion gene and also a leucaena cDNA encoding an enzyme for mimosine degradation expecting that the transformed plants will contain reduced amounts of mimosine.

Transcriptome analysis of Acacia koa.  Acacia koa (koa) is a important timber wood tree in Hawaii.  It is a leguminous tree and previously one of my students, James Leary, discovered canopy nodulation from the adventitous roots of koa. In recent years, wilt and dyeback diseases, caused by Fusarium oxysporum f. sp. koae, have caused heavy damage to the native koa forests in the Hawaiian Islands.  As a cross-pollinated tree species, selection for true-breeing wilt-resistant koa is not possible.  In collaboration with Nick Dudley at the Hawaii Agriculture Research center, we have developed a greenhouse screening method for selection of wilt-resistant koa seedlings and families.  We are now working on transcriptome analysis of koa with the aim to identify genes for disease resistance and wood development.

Recent publications

Negi VS, Bingham J-P, Li QX, Borthakur D (2013) midD-encoded ‘rhizomimosinase’ from Rhizobium sp. strain TAL1145 is a C–N lyase that catabolizes L-mimosine into 3-hydroxy-4-pyridone, pyruvate and ammonia. Amino Acids (in press).

Adamski DJ, Dudley NS, Morden CW, Borthakur D (2013) Cross-amplification of non-native Acacia species in the Hawaiian Islands using microsatellite markers from Acacia koa.  Plant Biosystems 146:  (in press)

Pal A, Negi VS, Khanal S, Borthakur D (2012) Immunodetection of curcin in seed meal of Jatropha curcas using polyclonal antibody developed against curcin-L. Current Nutrition & Food Science 8: 213-219.

Rushanaedy I, Jones TC, Dudley NS, Liao RJF, Agbayani R, Borthakur D (2012) Chitinase is a potential molecular biomarker for detecting resistance to Fusarium oxysporum in Acacia koa.  Tropical Plant Biol. 5:244–252.  DOI 10.1007/s12042-012-9108-7.

Adamski DJ, Dudley NS, Morden CW, Borthakur D (2012) Genetic differentiation and diversity of Acacia koa populations in the Hawaiian Islands.  Plant Species Biology. 27: 181-190 (with cover page photo from our work) DOI: 10.1111/j.1442-1984.2011.00359.x

Pal A, Negi VS, Borthakur D (2012) Efficient in vitro regeneration of Leucaena leucocephala using immature zygotic embryos as explants. Agroforestry Systems 84:131–140  (DOI 10.1007/s10457-011-9438-8).

Singh VS, Pal A, Singh R, Borthakur D (2011) Identification of species-specific genes from Leucaena leucocephala using interspecies suppression subtractive hybridization.  Annals of applied Biology 159: 387–398  (doi:10.1111/j.1744-7348.2011.00506.x).

Walton CB, Jube S, Schrlemmer A, Patek PQ, Zimmerman DH, Rosenthal KS, Borthakur D (2010) Ex vivo stimulation assay for T-cell responses for tuberculosis using LEAPS-peptide heteroconjugates. Current Trends in Microbiology 6:1-12.

Jube SLR, Borthakur D (2010) Transgenic Leucaena leucocephala expressing the Rhizobium gene pydA encoding a meta-cleavage dioxygenase shows reduced mimosine content. Plant Physiol Biochem 48 (2010) 273-278

Jube S, Awaya J, Borthakur D (2009) Expression of Rhizobium pydA-pydB fusion gene in Nicotiana tabacum confers resistance to the toxic aromatic compound 3-hydroxy-4-pyridone. Biologia Plantarum 53 (2): 355-359, 2009

Jube S, Borthakur D (2009) Development of an Agrobacterium-mediated transformation protocol for the recalcitrant tree-legume Leucaena leucocephala using immature zygotic embryos. Plant Cell, Tissue and Organ Culture (PCTOC): Journal of Plant Biotechnology 96: 325-333.

Fredua-Agyeman R, Adamski D, Liao RJ, Morden C, Borthakur D (2008) Development and characterization of microsatellite markers for analysis of population differentiation in the tree legume Acacia koa (Fabaceae: Mimosoideae) in the Hawaiian Islands.  Genome 51: 1001-1015.

Walton CB, Inos ABH, Andres OA, Jube S, de Couet HG, Douglas JT, Patek PQ, Borthakur D (2008) Immunization with hybrid recombinant Mycobacterium tuberculosis H37Rv proteins increases the TH1 cytokine response in mice following a pulmonary instillation of irradiated mycobacteria.  Vaccine 26 26,4396-4402.

Kutin RK, Jenkins DM, Borthakur D (2008) Characterization of a Corynebacterium strain that can reduce nitrate from high strength nitrate medium. Bioremediation Journal. 12(3):168-172.

Awaya JD, Tittabutr P, Li QX, Borthakur D (2008) Pyruvate carboxylase is involved in metabolism of mimosine by Rhizobium sp. strain TAL1145. Archives of Microbiology.  DOI:  10.1007/s00203-008-0384-4

Tittabutr P, Awaya JD, Li QX, Borthakur D (2008) The cloned 1-aminocyclopropane-1-carboxylate (ACC) deaminase gene from Sinorhizobium sp. strain BL3 in Rhizobium sp. strain TAL1145 promotes nodulation and growth of Leucaena leucocephala. Systematic and Applied Microbiology 31:141-150.

Teaching

• MBBE401/ Biol401 Molecular Biotechnology (3 cr) spring semester

• MBBE601/ Micr601 Molecular Cell Biology (3 cr) fall semester

• MBBE375 Multidisciplinary Biochemistry (3 cr) fall semester

Other academic activities

• Chairman, Graduate Program in Molecular Biosciences and Bioengineering, University of Hawaii (1996 - present).

• Member of the Editorial Board, Applied and Environmental Microbiology (2001-2009).

• Member of the Editorial Board and one of the Editors, International Journal of Microbiology (2008 -present)

• Member of the Editorial Board and Manuscript Editor, Symbiosis

• Assistant Editor, World Journal of Microbiology & Biotechnology, Kluwer Academic Publishers, Dordrecht, The Netherlands (1999-2004).

• Regular reviewer for ‘Soil Biology & Biochemistry’, ad hoc reviewer for Microbiology’, ‘Archives of Microbiology’ and ‘Journal of Bacteriology’.

• Grant review panel member for National Science Foundation (2010), USDA-NIFA (2012)

• Member of the Organizing Committee, CTAHR Symposium (2001 -present)

Awards

• 2004 'North American Colleges and Teachers of Agriculture Teaching award of Merit'.

• 2004 'CTAHR Excellence in Teaching' Award.

• 2003 'Most Motivational CTAHR Faculty' award by the College of Tropical Agriculture and Human Resources, University of Hawaii at Manoa.

Award received by my students

• Jannai Yafuso: CTAHR Best MS Student Oral Presentation $1500 Travel Award in the 2012 CTAHR Symposium.

• Vishal Negi: Gamma Sigma Delta PhD Student Oral Presentation Award in the 2012 CTAHR Symposium.

• Jannai Yafuso: Second place in MS Poster Presentation in the 2011 ASM Hawaii Symposium.

• Vishal Negi: CTAHR PhD Student Oral Presentation Award of Merit in the 2011 CTAHR Symposium.

• Jannai Yafuso: CTAHR MS Student Oral Presentation Award of Merit in the 2011 CTAHR Symposium.

• Vishal Negi: Second place in Oral Presentation $300 Award the 2010 ASM Hawaii Symposium.

• Tyler Jones: CTAHR Best MS Student Oral Presentation $1500 Travel Award in the 2010 CTAHR Symposium.

• Vishal Negi: Gamma Sigma Delta PhD Student Oral Presentation Award of Merit Vishal Negi in the 2010 CTAHR Symposium.

• Kehaulani Lee: Best Undergraduate Oral Presentation Award in the 2009 CTAHR Symposium.

• Sandro Jube: CTAHR Best PhD Student Poster Presentation $1500 Travel Award in the 2008 CTAHR Symposium.

• Daniel Adamski: CTAHR Best MS Student Oral Presentation $1500 Travel Award in the 2008 CTAHR symposium.

• Rudolph Fredua-Agyeman: Best PhD poster presentation among MBBE students in the 2007 CTAHR Symposium

• Sandro Jube: CTAHR Best PhD Student Oral Presentation $1500 Travel Award in the 2007 CTAHR Symposium.

• Rudolph Fredua-Agyeman: MBBE Best PhD student poster presentation in the 2007 CTAHR Symposium

• Dan Adamski: CTAHR Best MS Student poster Presentation $1500 Travel Award in the 2007 CTAHR Symposium

• Chad Walton: Case Foundation Business Plan Award $16,000 (2006)

• Jon Awaya: National Science Foundation Postdoctoral Award (2006) to conduct postdoctoral research at University of Notre Dame

• Sandro Jube: First prize in MS oral presentation in the 2007 CTAHR symposium in the 2005 CTAHR symposium.

• James Leary: Second prize in PhD oral presentation in the 2005 CTAHR symposium.

• Jon Awaya: Gamma Sigma Delta Doctoral Student Oral Presentation Award of Merit 2005.

• Jon Awaya: First prize for oral presentation, American Society of Microbiology, Hawaii 2005.

• Chad Walton: 2005 INBRE Research Award.

• Chad Walton: 2005 BRIN Research Award.

• Jon Awaya: ARC Foundation Excellence in Research Award 2004.

• Jon Awaya: Gamma Sigma Delta Doctoral Student Oral Presentation Award of Merit 2004.

• Jon Awaya: First prize for oral presentation, American Society of Microbiology, Hawaii 2004.

• Chad Walton: 2004 BRIN Research Award.

• Chad Walton: MBBE Departmental Award for oral presentation, CTAHR symposium 2004.

• James Leary: ARC Foundation Excellence in Research Award 2003.

• James Leary: CTAHR Best PhD Student Oral Presentation $1500 Travel Award in the 2003 CTAHR symposium.

• Jon Awaya: Gamma Sigma Delta Doctoral Student Oral Presentation Award of Merit 2003.

• Jon Awaya: Best in MBBE oral presentation in the 2003 CTAHR symposium.

• Havila Saafi: Best in MBBE poster presentation in the 2003 CTAHR symposium.

• Jon Awaya: First prize in the research symposium organized by the American Society for Microbiology Hawaii Chapter on April 5, 2003.

• Chad Walton: First prize in the Biomedical Research Symposium organized by the John Burn Medical School on April 25, 2003.

• Ana Blossom Inos: First prize for oral presentation in the Annual Biomedical Research Conference for Minority Students held in San Diego, in October 15-18, 2003.

• Paul Fox: First prize in the Biomedical Research Symposium organized by the John Burn Medical School, 2002.

• Nikhat Parveen:  Gamma Sigma Delta PhD Student Oral Presentation Award of Merit in 1995 CTAHR Symposium

Current graduate students

1. Archana Pal (2009-  ) PhD. Dissertation subject: The effects of cytosolic and chloroplastic carbonic anhydrases on stress tolerance in Leucaena leucocephala. 

2. Michael Honda (2012- ) MS. Thesis title: Expression of mimosinase in Leucaena leucocephala.

3. Dung Pham (2011 - ) MS. Thesis title: Identification of stress-related genes in Leucaena leucocephala through transcriptome analysis.

4. Jadd Correia (2012- ) MS. Thesis title: Genetic transformation of Leucaena leucocephala.

5. Kazue Ishihara (2011- ) MS. Thesis title: Transcriptome analysis in Acacia koa – identification of genes for wood formation.

6. Jannai Yafuso (2010- ) MS. Thesis title: Cysteine and mimosine biosynthesis catalyzed by cysteine/mimosine synthase from Leucaena leucocephala.

Past students

PhD:

1. Vishal Negi (2007-2012) PhD.  Dissertation title:  Biochemistry of mimosine catabolism by enzymes from Rhizobium sp. TAL1145 and Leucaena leucocephala.

2. Sandro Jube (2005-2009) PhD. Dissertation subject: Genetic transformation of Leucaena leucocephala.

3. Rudolph Fredua-Agyeman (2005-2008) PhD. Dissertation subject: Genetic diversity in Acacia koa in Hawaiian Islands. 

4. Chad Walton (2002-06) PhD.  Dissertation subject: Recombinant hyrbrid vaccine development against Mycobacterium tuberculosis..

5. James Leary (2001-07) Ph.D. Dissertation subject: Bradyrhizobium inoculant of Acacia koa.

6. Jonathan Awaya (2001-05) Ph.D. Dissertation subject: Analysis of pyd gene cluster in Rhizobium.

7. Pakieli Kafusi (1999-05) Ph.D. dissertation: Regulation of exopolysaccharide synthesis in Rhizobium by an alternate sigma factor.

8. Paul Fox (1999-02) Ph.D. dissertation: Biochemistry and genetics of mimosine degradation in Rhizobium.

9. Rongguan Jin (1995-1999) Ph.D. dissertation: Genetic engineering of cabbage and watercress with B.T. cry genes for insect resistance.

10. Muchdar Soedarjo (1993-1997) Ph.D. dissertation: Genetics of mimosine degradation by Rhizobium sp. strain TAL1145 that nodulates Leucaena.   

11. Nikhat Parveen (1991-1995) Ph.D. dissertation: Genetics of exopolysaccharide synthesis in Rhizobium sp. strain TAL1145 that nodulates tree legumes.

MS (with thesis):

1. Daniel Adamski (2006-08) MS.  Thesis title:  Molecular phylogenetics and diversity of the Acacia koa complex based on DNA sequences and microsatellite markers.

2. Masaki Nasu (2003-06) MS.  Thesis subject: Developing molecular markers for detection of homozygous lyretail fin type in sword fish.

3. Sandro Jube (2002-05) MS. Thesis subject.  Expression of Rhizobium dioxygenase and hydrolase genes in tobacco.

4. Joy Anamizu (1999-02) M.S. thesis:  Isolation, identification and characterization of mimosine-degrading soil bacteria isolated from the rhizosphere of Leucaena leucocephala.

5. Havila Saafi (2000-01) M.S.  thesis: Tissue culture of Leucaena leucocephala.

6. Zerong You (1996-1998) M.S. Thesis: A stomatin-like protein from Rhizobium etli is required for nodulation competition on Phaseolus bean.

7. Jyothirmai Gubili (1995-1997) M.S. thesis: Isolation and characterization of uptake hydrogenase  genes from Anabaena sp. strain PCC7120.

8.  Xuefeng Gao (1992-1995) M.S. thesis:  Genetic analysis of nodulation competitiveness in Rhizobium  etli.

9.  Shahriar Pooyan (1990-1992): MS thesis: Identification and characterization of chromosomal genes in Rhizobium  required for nitrogen fixation in beans.

MS (plan-B)

1. Tomomi Ida MS 2012. Characterization of a gene for stress tolerance from Leucaena leucocephala. (Advisor: Dulal Borthakur)

2. Eric K. W. Lee MS 2012. Tissue culture and genetic transformation of Leucaena leucocephala. (Advisor: Dulal Borthakur)

3. Asmahan Y. Abdallah MS. 2005. Recombinant vaccine against Mycobacterium tuberculosis. (Advisor: Dulal Borthakur)

4. Carie Tome MS 2002. Isolation of mimosine-degradation defective mutants of Rhizobium sp. strain TAL1145.  (Advisor: Dulal Borthakur)

5. Thomas Bui 1999. Construction of cDNA library Leucaena leucocephala. (Advisor: Dulal Borthakur)

6. Benjamin Tran 1998. Tissue culture of watercress. (Advisor: Dulal Borthakur)

BS (honors) with thesis

1. Ana Blossom Innos  BS (Honors) 2005. Title of thesis: Identification of Mycobacterium tuberculosis genes for recombinant vaccine development against tuberculosis (Chair: Dulal Borthakur)

2. Wendy Higashide BS (Honors) 2000 Characterization of Rhizobium gene involved in lipopolysaccharide synthesis. (Chair: Dulal Borthakur, Biology)