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Novel low-temperature-active, salt-tolerant and proteases-resistant endo-1,4-β-mannanase from a new Sphingomonas strain

Paper ID Volume ID Publish Year Pages File Format Full-Text
21135 43208 2012 7 PDF Available
Title
Novel low-temperature-active, salt-tolerant and proteases-resistant endo-1,4-β-mannanase from a new Sphingomonas strain
Abstract

Sphingomonas sp. JB13, isolated from slag of a > 20-year-old phosphate rock-stacking site, showed the highest 16S rDNA (1343 bp) identity of 97.2% with Sphingomonas sp. ERB1–3 (FJ948169) and < 97% identities with other identified Sphingomonas strains. A mannanase-coding gene (1191 bp) was cloned and encodes a 396-residue polypeptide (ManAJB13) showing the highest amino acid sequence identities of 56.2% with the putative glycosyl hydrolase (GH) family 26 endo-1,4-β-mannanase from Rhodothermus marinus (YP_004824245), and 44.2% with the identified GH 26 endo-1,4-β-mannanase from Cellvibrio japonicus (2VX5_A). The recombinant ManAJB13 (rManAJB13) was expressed in Escherichia coli BL21 (DE3). Purified rManAJB13 displayed the typical characteristics of low-temperature-active enzymes: showing apparent optimal at 40°C, ~ 55% of the maximum activity at 20°C and ~ 20% at 10°C, and thermolability at 45°C (~ 15 min half-life). The potential mechanism for low-temperature-activity of GH 26 endo-1,4-β-mannanases might be ascribed to the more hydrophobic residues (AILFWV) and less polar residues (NCQSTY) compared with typical thermophilic and mesophilic counterparts. The purified rManAJB13 exhibited > 85% mannanase activity at the concentration of 0–4.0 M NaCl. No loss of enzyme activity was observed after incubating the enzyme with 1 M or 2 M NaCl, or trypsin or proteinase K at 37°C and pH 6.5 for 1 h. The Km, Vmax and kcat values were 5.0 mg ml− 1, 277.8 μmol min− 1 mg− 1, and 211.9 s− 1, respectively, using locust bean gum as the substrate.

Keywords
Sphingomonas; Mannanase; Low-temperature activity; Salt tolerance; Proteases-resistance
First Page Preview
Novel low-temperature-active, salt-tolerant and proteases-resistant endo-1,4-β-mannanase from a new Sphingomonas strain
Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Bioscience and Bioengineering - Volume 113, Issue 5, May 2012, Pages 568–574
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering