Back to Search
Start Over
Recombinant expression and molecular characterization of buffalo sperm lysozyme-like protein 1.
- Source :
-
Protein expression and purification [Protein Expr Purif] 2022 Feb; Vol. 190, pp. 105993. Date of Electronic Publication: 2021 Oct 14. - Publication Year :
- 2022
-
Abstract
- Several sperm lysozyme-like genes evolved from lysozyme by successive duplications and mutations; however their functional role in the reproduction of farm animals is not well understood. To understand the function and molecular properties of buffalo sperm lysozyme-like protein 1 (buSLLP1), it was expressed in E. coli; however, it partitioned to inclusion bodies. Lowering of temperature and inducer concentration did not help in the recovery of the expressed protein in the biologically active form. Therefore, buSLLP1 was cloned and expressed in Pichiapink system based on auxotrophic Pichia pastoris in a labscale fermenter. The expressed protein was obtained in flow-through by using a 30 kDa ultrafiltration membrane followed by MonoQ anion exchange chromatography, resulting in a homogenous preparation of 40 mg recombinant buSLLP1 per liter of initial spent culture-supernatant. Circular dichroism spectroscopy showed that recombinant buSLLP1 possessed a native-like secondary structure. The recombinant buSLLP1 also showed thermal denaturation profile typical of folded globular proteins; however, the thermal stability was lower than the hen egg white lysozyme. Binding of buSLLP1 to chitin and zona pellucida of buffalo oocytes showed that the recombinant buSLLP1 possessed a competent binding pocket, therefore, the produced protein could be used to study its functional role in the reproduction of farm animals.<br /> (Copyright © 2021 Elsevier Inc. All rights reserved.)
- Subjects :
- Animals
Escherichia coli genetics
Escherichia coli metabolism
Recombinant Proteins biosynthesis
Recombinant Proteins chemistry
Recombinant Proteins genetics
Recombinant Proteins isolation & purification
Saccharomycetales genetics
Saccharomycetales metabolism
Buffaloes genetics
Gene Expression
Muramidase biosynthesis
Muramidase chemistry
Muramidase genetics
Muramidase isolation & purification
Subjects
Details
- Language :
- English
- ISSN :
- 1096-0279
- Volume :
- 190
- Database :
- MEDLINE
- Journal :
- Protein expression and purification
- Publication Type :
- Academic Journal
- Accession number :
- 34656738
- Full Text :
- https://doi.org/10.1016/j.pep.2021.105993