Probing C-terminal interactions of the Pseudomonas stutzeri cyanide-degrading CynD protein

 

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dc.contributor.author Crum, Mary Abou-Nader
dc.contributor.author Park, Jason M
dc.contributor.author Mulelu, Andani E
dc.contributor.author Sewell, Trevor B
dc.contributor.author Benedik, Michael J
dc.date.accessioned 2016-09-02T11:04:40Z
dc.date.available 2016-09-02T11:04:40Z
dc.date.issued 2015
dc.identifier http://dx.doi.org/10.1007/s00253-014-6335-x
dc.identifier.citation Crum, M. A. N., Park, J. M., Mulelu, A. E., Sewell, B. T., & Benedik, M. J. (2015). Probing C-terminal interactions of the Pseudomonas stutzeri cyanide-degrading CynD protein. Applied microbiology and biotechnology, 99(7), 3093-3102. en_ZA
dc.identifier.issn 0175-7598 en_ZA
dc.identifier.uri http://hdl.handle.net/11427/21658
dc.identifier.uri http://link.springer.com/article/10.1007/s00253-014-6335-x
dc.description.abstract The cyanide dihydratases from Bacillus pumilus and Pseudomonas stutzeri share high amino acid sequence similarity throughout except for their highly divergent C-termini. However, deletion or exchange of the C-termini had different effects upon each enzyme. Here we extended previous studies and investigated how the C-terminus affects the activity and stability of three nitrilases, the cyanide dihydratases from B. pumilus (CynDpum) and P. stutzeri (CynDstut) and the cyanide hydratase from Neurospora crassa. Enzymes in which the C-terminal residues were deleted decreased in both activity and thermostability with increasing deletion lengths. However, CynDstut was more sensitive to such truncation than the other two enzymes. A domain of the P. stutzeri CynDstut C-terminus not found in the other enzymes, 306GERDST311, was shown to be necessary for functionality and explains the inactivity of the previously described CynDstut-pum hybrid. This suggests that the B. pumilus C-terminus, which lacks this motif, may have specific interactions elsewhere in the protein, preventing it from acting in trans on a heterologous CynD protein. We identify the dimerization interface A-surface region 195–206 (A2) from CynDpum as this interaction site. However, this A2 region did not rescue activity in C-terminally truncated CynDstutΔ302 or enhance the activity of full-length CynDstut and therefore does not act as a general stability motif. en_ZA
dc.language eng en_ZA
dc.publisher Springer Verlag en_ZA
dc.source Applied Microbiology and Biotechnology en_ZA
dc.source.uri http://link.springer.com/journal/253
dc.subject.other Cyanide
dc.subject.other dihydratase
dc.subject.other Nitrilase
dc.subject.other Cyanide
dc.subject.other Bioremediation
dc.title Probing C-terminal interactions of the Pseudomonas stutzeri cyanide-degrading CynD protein en_ZA
dc.type Journal Article en_ZA
dc.date.updated 2016-09-02T11:03:41Z
uct.type.publication Research en_ZA
uct.type.resource Article en_ZA
dc.publisher.institution University of Cape Town
uct.type.filetype Text
uct.type.filetype Image
dc.identifier.apacitation Crum, M. A., Park, J. M., Mulelu, A. E., Sewell, T. B., & Benedik, M. J. (2015). Probing C-terminal interactions of the Pseudomonas stutzeri cyanide-degrading CynD protein. <i>Applied Microbiology and Biotechnology</i>, http://hdl.handle.net/11427/21658 en_ZA
dc.identifier.chicagocitation Crum, Mary Abou-Nader, Jason M Park, Andani E Mulelu, Trevor B Sewell, and Michael J Benedik "Probing C-terminal interactions of the Pseudomonas stutzeri cyanide-degrading CynD protein." <i>Applied Microbiology and Biotechnology</i> (2015) http://hdl.handle.net/11427/21658 en_ZA
dc.identifier.vancouvercitation Crum MA, Park JM, Mulelu AE, Sewell TB, Benedik MJ. Probing C-terminal interactions of the Pseudomonas stutzeri cyanide-degrading CynD protein. Applied Microbiology and Biotechnology. 2015; http://hdl.handle.net/11427/21658. en_ZA
dc.identifier.ris TY - Journal Article AU - Crum, Mary Abou-Nader AU - Park, Jason M AU - Mulelu, Andani E AU - Sewell, Trevor B AU - Benedik, Michael J AB - The cyanide dihydratases from Bacillus pumilus and Pseudomonas stutzeri share high amino acid sequence similarity throughout except for their highly divergent C-termini. However, deletion or exchange of the C-termini had different effects upon each enzyme. Here we extended previous studies and investigated how the C-terminus affects the activity and stability of three nitrilases, the cyanide dihydratases from B. pumilus (CynDpum) and P. stutzeri (CynDstut) and the cyanide hydratase from Neurospora crassa. Enzymes in which the C-terminal residues were deleted decreased in both activity and thermostability with increasing deletion lengths. However, CynDstut was more sensitive to such truncation than the other two enzymes. A domain of the P. stutzeri CynDstut C-terminus not found in the other enzymes, 306GERDST311, was shown to be necessary for functionality and explains the inactivity of the previously described CynDstut-pum hybrid. This suggests that the B. pumilus C-terminus, which lacks this motif, may have specific interactions elsewhere in the protein, preventing it from acting in trans on a heterologous CynD protein. We identify the dimerization interface A-surface region 195–206 (A2) from CynDpum as this interaction site. However, this A2 region did not rescue activity in C-terminally truncated CynDstutΔ302 or enhance the activity of full-length CynDstut and therefore does not act as a general stability motif. DA - 2015 DB - OpenUCT DP - University of Cape Town J1 - Applied Microbiology and Biotechnology LK - https://open.uct.ac.za PB - University of Cape Town PY - 2015 SM - 0175-7598 T1 - Probing C-terminal interactions of the Pseudomonas stutzeri cyanide-degrading CynD protein TI - Probing C-terminal interactions of the Pseudomonas stutzeri cyanide-degrading CynD protein UR - http://hdl.handle.net/11427/21658 ER - en_ZA


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