DOI: 10.1111/j.1432-1033.2004.03988.x. Identification of a gene encoding Lon protease from <i>Brevibacillus thermoruber</i> WR‐249 and biochemical characterization of its thermostable recombinant enzyme

Identification of a gene encoding Lon protease from Brevibacillus thermoruber WR‐249 and biochemical characterization of its thermostable recombinant enzyme

10.1111/j.1432-1033.2004.03988.x

Crossref journal-article

Wiley

European Journal of Biochemistry (311)

Abstract

A gene encoding thermostable Lon protease from Brevibacillus thermoruber WR‐249 was cloned and characterized. The Br. thermoruber Lon gene (Bt‐lon) encodes an 88 kDa protein characterized by an N‐terminal domain, a central ATPase domain which includes an SSD (sensor‐ and substrate‐discrimination) domain, and a C‐terminal protease domain. The Bt‐lon is a heat‐inducible gene and may be controlled under a putative Bacillus subtilisσA‐dependent promoter, but in the absence of CIRCE (controlling inverted repeat of chaperone expression). Bt‐lon was expressed in Escherichia coli, and its protein product was purified. The native recombinant Br. thermoruber Lon protease (Bt‐Lon) displayed a hexameric structure. The optimal temperature of ATPase activity for Bt‐Lon was 70 °C, and the optimal temperature of peptidase and DNA‐binding activities was 50 °C. This implies that the functions of Lon protease in thermophilic bacteria may be switched, depending on temperature, to regulate their physiological needs. The peptidase activity of Bt‐Lon increases substantially in the presence of ATP. Furthermore, the substrate specificity of Bt‐Lon is different from that of E. coli Lon in using fluorogenic peptides as substrates. Notably, the Bt‐Lon protein shows chaperone‐like activity by preventing aggregation of denatured insulin B‐chain in a dose‐dependent and ATP‐independent manner. In thermal denaturation experiments, Bt‐Lon was found to display an indicator of thermostability value, Tm of 71.5 °C. Sequence comparison with mesophilic Lon proteases shows differences in the rigidity, electrostatic interactions, and hydrogen bonding of Bt‐Lon relevant to thermostability.

Bibliography

Lee Tsay, A. Y. âL., SanâSan, Chen, M., & Wu, S. (2004). Identification of a gene encoding Lon protease from Brevibacillus thermoruber WRâ249 and biochemical characterization of its thermostable recombinant enzyme. European Journal of Biochemistry, 271(4), 834â844. Portico.

Authors 3

Alan Y.‐L., San‐San Lee Tsay (first)
Mao‐Yen Chen (additional)
Shih‐Hsiung Wu (additional)

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Dates

Type	When
Created	21 years, 7 months ago (Feb. 5, 2004, 7:01 a.m.)
Deposited	1 year, 10 months ago (Oct. 31, 2023, 12:11 p.m.)
Indexed	1 year, 10 months ago (Nov. 1, 2023, 3:35 a.m.)
Issued	21 years, 7 months ago (Feb. 1, 2004)
Published	21 years, 7 months ago (Feb. 1, 2004)
Published Online	21 years, 7 months ago (Feb. 2, 2004)
Published Print	21 years, 7 months ago (Feb. 1, 2004)

Funders 0

None

BibTeX

@article{Lee_Tsay_2004, title={Identification of a gene encoding Lon protease from Brevibacillus thermoruber WR‐249 and biochemical characterization of its thermostable recombinant enzyme}, volume={271}, ISSN={1432-1033}, url={http://dx.doi.org/10.1111/j.1432-1033.2004.03988.x}, DOI={10.1111/j.1432-1033.2004.03988.x}, number={4}, journal={European Journal of Biochemistry}, publisher={Wiley}, author={Lee Tsay, Alan Y.‐L., San‐San and Chen, Mao‐Yen and Wu, Shih‐Hsiung}, year={2004}, month=feb, pages={834–844} }

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