Open Access

Biosynthesis, purification, and biochemical properties of an alkaline-thermostable keratinase derived from Bacillus cereus J6

Rongxian Zhang

Rongxian Zhang

  • School of Ecological Engineering, Guizhou University of Engineering ScienceBijie, China
  • Key Laboratory of Biological Resources Development and Ecological Restoration in Guizhou ProvinceBijie, China
Search for this author on
Sciendo|Google Scholar
Zhang, Rongxian
, 
Chengguo Fu

Chengguo Fu

  • Key Laboratory of Biological Resources Development and Ecological Restoration in Guizhou ProvinceBijie, China
  • School of Mechanical Engineering, Guizhou University of Engineering ScienceBijie, China
Search for this author on
Sciendo|Google Scholar
Fu, Chengguo
, 
Yipeng Feng

Yipeng Feng

Mechatronic Engineering and Intelligent Manufacturing College, Henan Open UniversityZhengzhou, China
Search for this author on
Sciendo|Google Scholar
Feng, Yipeng
, 
Bian Hu

Bian Hu

  • School of Ecological Engineering, Guizhou University of Engineering ScienceBijie, China
  • Key Laboratory of Biological Resources Development and Ecological Restoration in Guizhou ProvinceBijie, China
Search for this author on
Sciendo|Google Scholar
Hu, Bian
, 
Yizhong Zhang

Yizhong Zhang

  • School of Ecological Engineering, Guizhou University of Engineering ScienceBijie, China
  • Key Laboratory of Biological Resources Development and Ecological Restoration in Guizhou ProvinceBijie, China
Search for this author on
Sciendo|Google Scholar
Zhang, Yizhong
, 
Wei Zhang

Wei Zhang

  • School of Ecological Engineering, Guizhou University of Engineering ScienceBijie, China
  • Key Laboratory of Biological Resources Development and Ecological Restoration in Guizhou ProvinceBijie, China
Search for this author on
Sciendo|Google Scholar
Zhang, Wei
 and 
Qunying Xiao

Qunying Xiao

  • School of Ecological Engineering, Guizhou University of Engineering ScienceBijie, China
  • Key Laboratory of Biological Resources Development and Ecological Restoration in Guizhou ProvinceBijie, China
Search for this author on
Sciendo|Google Scholar
Xiao, Qunying
  
Mar 17, 2025
Biosynthesis, purification, and biochemical properties of an alkaline-thermostable keratinase derived from Bacillus cereus J6's Cover Image
About this article
Previous Article
Next Article
Cite
Download Cover
Published Online: Mar 17, 2025
Received: Oct 30, 2024
Accepted: Feb 10, 2025
DOI: https://doi.org/10.2478/amns-2025-0831
Keywords
© 2025 Rongxian Zhang et al., published by Sciendo
This work is licensed under the Creative Commons Attribution 4.0 International License.

Figure 1.

Phylogenetic tree of Strain J6 constructed via Neighbor-joining method
Phylogenetic tree of Strain J6 constructed via Neighbor-joining method

Figure 2.

Keratinase production by B. cereus J6. A. Enzyme production profile; B, Feather; C, Nitrogen sources; D, Urea; E, Carbon sources; F, Lactose; G, Mineral salts; F, MgSO4.
Keratinase production by B. cereus J6. A. Enzyme production profile; B, Feather; C, Nitrogen sources; D, Urea; E, Carbon sources; F, Lactose; G, Mineral salts; F, MgSO4.

Figure 3.

Impact of cultural conditions on B. cereus J6 producing keratinase
Impact of cultural conditions on B. cereus J6 producing keratinase

Figure 4.

SDS-PAGE analysis of the purification of J6 keratinase. Lane M, protein marker; Lane 1, the crude B. cereus J6 keratinase; Lane 2, ASP; Lane 3, the target enzyme.
SDS-PAGE analysis of the purification of J6 keratinase. Lane M, protein marker; Lane 1, the crude B. cereus J6 keratinase; Lane 2, ASP; Lane 3, the target enzyme.

Figure 5.

Impact of temperature and pH
Impact of temperature and pH

Figure 6.

The enzymatic activity of the keratinase towards various substrates
The enzymatic activity of the keratinase towards various substrates

Figure 7.

The kinetics of the purified keratinase
The kinetics of the purified keratinase

Effects of chemicals on J6 keratinase

Chemicals Concentration Relative activity (%)
Na+ 5 mmol/L 102.59±2.78
K+ 5 mmol/L 99.03±2.93
Mn2+ 5 mmol/L 106.73±2.25
Ca2+ 5 mmol/L 122.56±2.19
Mg2+ 5 mmol/L 131.81±5.14
Zn2+ 5 mmol/L 118.52±3.52
Cu2+ 5 mmol/L 60.87±4.53
Al3+ 5 mmol/L 62.67±6.21
Fe2+ 5 mmol/L 52.89±5.35
Fe3+ 5 mmol/L 31.04±3.87
Tween 20 1% 115.67±3.14
Tween 60 1% 175.27±1.31
Tween 80 1% 150.16±2.31
Triton X-100 1% 106.06±0.52
SDS 1% 84.30±2.56
DTT 1 mmol/L 155.96±2.73
EDTA 1 mmol/L 5.05±2.21
PMSF 1 mmol/L Not detected

Results of orthogonal experiment

Number A B C D Keratinase yield (U/mL)
1 1 1 1 1 375.55±3.12
2 1 2 2 2 179.8±5.93
3 1 3 3 3 359.9±6.49
4 2 1 2 3 401.4±9.84
5 2 2 3 1 342.55±3.16
6 2 3 1 2 208.9±9.53
7 3 1 3 2 186.9±42
8 3 2 1 3 307.25±4.78
9 3 3 2 1 294.9±2.65
K1 305.08 321.28 297.23 337.67
K2 317.62 276.53 292.03 192.86
K3 263.02 288.9 296.45 356.18
R 54.6 32.38 5.2 163.32
Order of priority: D > A > B > C
Optimal Combination: A2B1C1D3

Orthogonal experimental design of four factors and three levels

Level Factor
A feather (g/L) B Lactose (g/L) C Urea (g/L) D MgSO4 (g/L)
1 7.5 2.5 7.5 0.9
2 10 5 10 1
3 12.5 7.5 12.5 1.1

Purification of the keratinase produced by B_ cereus J6

Procedures Protein quantity (mg) activity (U) Specific activity (U/mg) Purification fold Recovery rate (%)
Supernatant 53.25 24032 451.31 1.00 100
ASP 15.41 14256 925.19 2.05 59.32
DEAE-650 M chromatography 0.65 2739 4215.24 9.34 11.36
Language:
English
Publication timeframe:
1 times per year
Journal Subjects:
Life Sciences, Life Sciences, other, Mathematics, Applied Mathematics, General Mathematics, Physics, Physics, other
Journal RSS Feed