One-step elimination of L-cysteine desulfhydrase from crude enzyme extracts of Pseudomonas sp. TS1138 using an immunomagnetic affinity matrix improves the enzymatic production of L-cysteine.
In this study, a high efficiency immunomagnetic affinity matrix was developed to eliminate L-cysteine desulfhydrase (CD), which decomposes L-cysteine, in crude enzyme extracts from Pseudomonas sp. TS1138. After cloning and expression in Escherichia coli, recombinant CD was purified to raise polyclonal antibodies from mice. The anti-CD antibody was cross-linked to staphylococcal protein A-magnetic cellulose microspheres (MCMS) with dimethyl pimelimidate (DMP). The natural CD was eliminated from the crude enzyme extracts by treatment with the cross-linked antibody-protein A-MCMS, resulting in a high level of L-cysteine production. The conversion rate of DL-2-amino-Delta2-thiazoline-4-carboxylic acid (DL-ATC) to L-cysteine increased significantly from 61.9 to 96.2%. The cross-linked antibody-protein A-MCMS showed its durability after repetitive use, maintaining a constant binding capacity for CD during five cycles. This study may lead to a convenient and cost-efficient method to produce L-cysteine by enzymatic conversions.
Yu
Yangsheng
Y
Department of Microbiology, College of Life Sciences, Nankai University, Tianjin 300071, China.
Bai
Gang
G
Liu
Chunqin
C
Cao
Yu
Y
Geng
Peng
P
Yang
Wenbo
W
D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D003542:Cystathionine gamma-Lyase / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D003545:Cysteine / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D008280:Magnetics; D008855:Microscopy, Electron, Scanning; D008863:Microspheres; D011549:Pseudomonas / Q000201:enzymology* / Q000235:genetics; D011773:Pyruvates / Q000378:metabolism; D011994:Recombinant Proteins / Q000276:immunology / Q000378:metabolism
D001426:Bacterial Proteins; D011773:Pyruvates; D011994:Recombinant Proteins; D003542:Cystathionine gamma-Lyase; D003545:Cysteine