Glutamyl endopeptidase from Enterococcus faecalis, designated SprE, is one of the important virulence factors secreted as zymogen. In the present study we expressed recombinant SprE proenzyme (pro-SprE) in Escherichia coli and investigated the in vitro processing to mature SprE. It was found that trypsin could efficiently produce the active form of SprE with the N-terminus Ser1 through cleavage between Arg-1 and Ser1 bond, which was subsequently auto-degraded into inactive species through the cleavage at the Glu6-Asp7 and Glu11-Val12 bonds. Although thermolysin could produce SprE with the N-terminus Leu2, but possessed no proteolytic activity. In contrast to the absolute requirement of the N-terminal Val1 in staphylococcal glutamyl endopeptidases, the N-terminal Ser1 of mature SprE could be substituted by other amino acids despite that Ser showed the maximal activity. Substitution of penultimate Leu2 of SprE to Val2 also reduced the activity to 40% of the wild type. Taken together, we conclude that pro-SprE was converted to mature form with the N-terminus Ser1 by a protease with specificity of trypsin and the length of the N-terminal region rather than specific residue is absolutely required for enzyme activity.
| Published in | Advances in Biochemistry (Volume 1, Issue 5) |
| DOI | 10.11648/j.ab.20130105.11 |
| Page(s) | 73-80 |
| Creative Commons |
This is an Open Access article, distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution and reproduction in any medium or format, provided the original work is properly cited. |
| Copyright |
Copyright © The Author(s), 2013. Published by Science Publishing Group |
Glutamyl Endopeptidase, Proenzyme Processing, Spre, Enterococcus faecalis, Gluv8
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APA Style
Shakh M. A. Rouf, Y. Ohara-Nemoto, T. Ono, Y. Shimoyama, S. Kimura, et al. (2013). In Vitro Processing of Glutamyl Endopeptidase Proenzymes from Enterococcus faecalis and Importance of N-terminal Residue in Enzyme Catalysis. Advances in Biochemistry, 1(5), 73-80. https://doi.org/10.11648/j.ab.20130105.11
ACS Style
Shakh M. A. Rouf; Y. Ohara-Nemoto; T. Ono; Y. Shimoyama; S. Kimura, et al. In Vitro Processing of Glutamyl Endopeptidase Proenzymes from Enterococcus faecalis and Importance of N-terminal Residue in Enzyme Catalysis. Adv. Biochem. 2013, 1(5), 73-80. doi: 10.11648/j.ab.20130105.11
AMA Style
Shakh M. A. Rouf, Y. Ohara-Nemoto, T. Ono, Y. Shimoyama, S. Kimura, et al. In Vitro Processing of Glutamyl Endopeptidase Proenzymes from Enterococcus faecalis and Importance of N-terminal Residue in Enzyme Catalysis. Adv Biochem. 2013;1(5):73-80. doi: 10.11648/j.ab.20130105.11
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Download@article{10.11648/j.ab.20130105.11,
author = {Shakh M. A. Rouf and Y. Ohara-Nemoto and T. Ono and Y. Shimoyama and S. Kimura and T. K. Nemoto},
title = {In Vitro Processing of Glutamyl Endopeptidase Proenzymes from Enterococcus faecalis and Importance of N-terminal Residue in Enzyme Catalysis},
journal = {Advances in Biochemistry},
volume = {1},
number = {5},
pages = {73-80},
doi = {10.11648/j.ab.20130105.11},
url = {https://doi.org/10.11648/j.ab.20130105.11},
eprint = {https://article.sciencepublishinggroup.com/pdf/10.11648.j.ab.20130105.11},
abstract = {Glutamyl endopeptidase from Enterococcus faecalis, designated SprE, is one of the important virulence factors secreted as zymogen. In the present study we expressed recombinant SprE proenzyme (pro-SprE) in Escherichia coli and investigated the in vitro processing to mature SprE. It was found that trypsin could efficiently produce the active form of SprE with the N-terminus Ser1 through cleavage between Arg-1 and Ser1 bond, which was subsequently auto-degraded into inactive species through the cleavage at the Glu6-Asp7 and Glu11-Val12 bonds. Although thermolysin could produce SprE with the N-terminus Leu2, but possessed no proteolytic activity. In contrast to the absolute requirement of the N-terminal Val1 in staphylococcal glutamyl endopeptidases, the N-terminal Ser1 of mature SprE could be substituted by other amino acids despite that Ser showed the maximal activity. Substitution of penultimate Leu2 of SprE to Val2 also reduced the activity to 40% of the wild type. Taken together, we conclude that pro-SprE was converted to mature form with the N-terminus Ser1 by a protease with specificity of trypsin and the length of the N-terminal region rather than specific residue is absolutely required for enzyme activity.},
year = {2013}
}
TY - JOUR T1 - In Vitro Processing of Glutamyl Endopeptidase Proenzymes from Enterococcus faecalis and Importance of N-terminal Residue in Enzyme Catalysis AU - Shakh M. A. Rouf AU - Y. Ohara-Nemoto AU - T. Ono AU - Y. Shimoyama AU - S. Kimura AU - T. K. Nemoto Y1 - 2013/12/30 PY - 2013 N1 - https://doi.org/10.11648/j.ab.20130105.11 DO - 10.11648/j.ab.20130105.11 T2 - Advances in Biochemistry JF - Advances in Biochemistry JO - Advances in Biochemistry SP - 73 EP - 80 PB - Science Publishing Group SN - 2329-0862 UR - https://doi.org/10.11648/j.ab.20130105.11 AB - Glutamyl endopeptidase from Enterococcus faecalis, designated SprE, is one of the important virulence factors secreted as zymogen. In the present study we expressed recombinant SprE proenzyme (pro-SprE) in Escherichia coli and investigated the in vitro processing to mature SprE. It was found that trypsin could efficiently produce the active form of SprE with the N-terminus Ser1 through cleavage between Arg-1 and Ser1 bond, which was subsequently auto-degraded into inactive species through the cleavage at the Glu6-Asp7 and Glu11-Val12 bonds. Although thermolysin could produce SprE with the N-terminus Leu2, but possessed no proteolytic activity. In contrast to the absolute requirement of the N-terminal Val1 in staphylococcal glutamyl endopeptidases, the N-terminal Ser1 of mature SprE could be substituted by other amino acids despite that Ser showed the maximal activity. Substitution of penultimate Leu2 of SprE to Val2 also reduced the activity to 40% of the wild type. Taken together, we conclude that pro-SprE was converted to mature form with the N-terminus Ser1 by a protease with specificity of trypsin and the length of the N-terminal region rather than specific residue is absolutely required for enzyme activity. VL - 1 IS - 5 ER -
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