Heterologous production of clavulanic acid intermediates in <Emphasis Type="Italic">Streptomyces venezuelae</Emphasis>

Heterologous expression can enhance production of diverse secondary metabolites by redirecting precursor pools towards compound of interest. In this study, Streptomyces venezuelae YJ028 was utilized as the heterologous host for the expression of four structural clavulanic acid biosynthesis genes, which encode carboxyethylarginine synthase (ceas2), β-lactam synthetase (bls2), clavaminate synthase (cas2), and proclavaminate amidinohydrolase (pah2). These genes were cloned into pIBR25 expression vector containing ermE* promoter to generate pBS4. The cas2 gene was also cloned into pSET152 to generate pCas2. It was then integrated into the genome of S. venezuelae YJ028. Upon metabolite profiling of recombinant strains by ultra-pressure liquid chromatography-photodiode array (UPLC-PDA) and high resolution liquid chromatography quadruple time-offlight electrospray ionization mass spectrometry (HR-LC-QTOF-ESI/MS), the production of following clavulanic acid intermediates in S. venezuelae recombinant were confirmed: deoxygaunidinoproclavaminic acid, guanidinoproclavaminic acid, and dihydroclavaminic acid. This work demonstrates the production of β-lactam intermediates of the clavulanic acid pathway by heterologous expression in S. venezuelae YJ028.