A synthetic microbial biosensor for high-throughput screening of lactam biocatalysts.

ABSTRACT: Biocatalytic cyclization is highly desirable for efficient synthesis of biologically derived chemical substances, such as the commodity chemicals ?-caprolactam and ?-valerolactam. To identify biocatalysts in lactam biosynthesis, we develop a caprolactam-detecting genetic enzyme screening system (CL-GESS). The Alcaligenes faecalis regulatory protein NitR is adopted for the highly specific detection of lactam compounds against lactam biosynthetic intermediates. We further systematically optimize the genetic components of the CL-GESS to enhance sensitivity, achieving 10-fold improvement. Using this highly sensitive GESS, we screen marine metagenomes and find an enzyme that cyclizes ?-amino fatty acids to lactam. Moreover, we determine the X-ray crystal structure and catalytic residues based on mutational analysis of the cyclase. The cyclase is also used as a helper enzyme to sense intracellular ?-amino fatty acids. We expect this simple and accurate biosensor to have wide-ranging applications in rapid screening of new lactam-synthesizing enzymes and metabolic engineering for lactam bio-production.

SUBMITTER: Yeom SJ 

PROVIDER: S-EPMC6265244 | biostudies-literature | 2018 Nov

REPOSITORIES: biostudies-literature