AUTHOR=Schaffert Lena , Schneiker-Bekel Susanne , Dymek Saskia , Droste Julian , Persicke Marcus , Busche Tobias , Brandt David , Pühler Alfred , Kalinowski Jörn 

TITLE=Essentiality of the Maltase AmlE in Maltose Utilization and Its Transcriptional Regulation by the Repressor AmlR in the Acarbose-Producing Bacterium Actinoplanes sp. SE50/110

JOURNAL=Frontiers in Microbiology

VOLUME=10

YEAR=2019

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2019.02448

DOI=10.3389/fmicb.2019.02448

ISSN=1664-302X

ABSTRACT=<p><italic>Actinoplanes</italic> sp. SE50/110 is the wild type of industrial production strains of the fine-chemical acarbose (acarviosyl-maltose), which is used as α-glucosidase inhibitor in the treatment of type II diabetes. Although maltose is an important building block of acarbose, the maltose/maltodextrin metabolism has not been studied in <italic>Actinoplanes</italic> sp. SE50/110 yet. Bioinformatic analysis located a putative maltase gene <italic>amlE</italic> (<italic>ACSP50_2474</italic>, previously named <italic>malL</italic>; <xref ref-type="bibr" rid="B74">Wendler et al., 2015a</xref>), in an operon with an upstream PurR/LacI-type transcriptional regulator gene, named <italic>amlR</italic> (<italic>ACSP50_2475</italic>), and a gene downstream (<italic>ACSP50_2473</italic>) encoding a GGDEF-EAL-domain-containing protein putatively involved in c-di-GMP signaling. Targeted gene deletion mutants of <italic>amlE</italic> and <italic>amlR</italic> were constructed by use of the CRISPR/Cas9 technology. By growth experiments and functional assays of Δ<italic>amlE</italic>, we could show that AmlE is essential for the maltose utilization in <italic>Actinoplanes</italic> sp. SE50/110. Neither a gene encoding a maltose phosphorylase (MalP) nor MalP enzyme activity were detected in the wild type. By this, the maltose/maltodextrin system appears to be fundamentally different from other described prokaryotic systems. By sequence similarity analysis and functional assays from the species <italic>Streptomyces lividans</italic> TK23, <italic>S. coelicolor</italic> A3(2) and <italic>S. glaucescens</italic> GLA.O, first hints for a widespread lack of MalP and presence of AmlE in the class Actinobacteria were given. Transcription of the <italic>aml</italic> operon is significantly repressed in the wild type when growing on glucose and repression is absent in an Δ<italic>amlR</italic> deletion mutant. Although AmlR apparently is a local transcriptional regulator of the <italic>aml</italic> operon, the Δ<italic>amlR</italic> strain shows severe growth inhibitions on glucose and – concomitantly – differential transcription of several genes of various functional classes. We ascribe these effects to <italic>ACSP50_2473</italic>, which is localized downstream of <italic>amlE</italic> and presumably involved in the metabolism of the second messenger c-di-GMP. It can be assumed, that maltose does not only represent the most important carbon source of <italic>Actinoplanes</italic> sp. SE50/110, but that its metabolism is coupled to the nucleotide messenger system of c-di-GMP.</p>