Improvement of Heterologous Soluble Expression of L-amino Acid Oxidase Using Logistic Regression.
Chembiochem
; 25(13): e202400243, 2024 Jul 02.
Article
in En
| MEDLINE
| ID: mdl-38696752
ABSTRACT
Successful implementation of enzymes in practical application hinges on the development of efficient mass production techniques. However, in a heterologous expression system, the protein is often unable to fold correctly and, thus, forms inclusion bodies, resulting in the loss of its original activity. In this study, we present a new and more accurate model for predicting amino acids associated with an increased L-amino acid oxidase (LAO) solubility. Expressing LAO from Rhizoctonia solani in Escherichia coli and combining random mutagenesis and statistical logistic regression, we modified 108 amino acid residues by substituting hydrophobic amino acids with serine and hydrophilic amino acids with alanine. Our results indicated that specific mutations in Euclidean distance, glycine, methionine, and secondary structure increased LAO expression. Furthermore, repeated mutations were performed for LAO based on logistic regression models. The mutated LAO displayed a significantly increased solubility, with the 6-point and 58-point mutants showing a 2.64- and 4.22-fold increase, respectively, compared with WT-LAO. Ultimately, using recombinant LAO in the biotransformation of α-keto acids indicates its great potential as a biocatalyst in industrial production.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Solubility
/
Escherichia coli
/
L-Amino Acid Oxidase
Language:
En
Journal:
Chembiochem
Journal subject:
BIOQUIMICA
Year:
2024
Document type:
Article
Affiliation country:
Japan