Ecker, A. K. | bioRxiv Science

bioRxiv2023

Structure and Biosynthesis of Hectoramide B, a Linear Depsipeptide from the Marine Cyanobacterium Moorena producens JHB Discovered via Co-culture with Candida albicans

The tropical marine cyanobacterium Moorena producens JHB is a prolific source of secondary metabolites with potential biomedical utility. Previous studies of this strain led to the discovery of several novel compounds such as the hectochlorins and jamaicamides; however, bioinformatic analyses of its genome suggested that there were many more cryptic biosynthetic gene clusters yet to be characterized. To potentially stimulate the production of novel compounds from this strain, it was co-cultured with Candida albicans. From this experiment, we observed the increased production of a new compound that we characterize here as hectoramide B. Bioinformatic analysis of the M. producens JHB genome enabled the identification of a putative biosynthetic gene cluster responsible for hectoramide B biosynthesis. This work demonstrates that co-culture competition experiments can be a valuable method to facilitate the discovery of novel natural products from cyanobacteria. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=106 SRC="FIGDIR/small/547815v1_ufig1.gif" ALT="Figure 1"> View larger version (19K): org.highwire.dtl.DTLVardef@1b507d4org.highwire.dtl.DTLVardef@152376org.highwire.dtl.DTLVardef@1cb410dorg.highwire.dtl.DTLVardef@11bcabc_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2023

Structural characterization of ligand binding and pH-specific enzymatic activity of mouse Acidic Mammalian Chitinase

Chitin is an abundant biopolymer and pathogen-associated molecular pattern that stimulates a host innate immune response. Mammals express chitin-binding and chitin-degrading proteins to remove chitin from the body. One of these proteins, Acidic Mammalian Chitinase (AMCase), is an enzyme known for its ability to function under acidic conditions in the stomach but is also active in tissues with more neutral pHs, such as the lung. Here, we used a combination of biochemical, structural, and computational modeling approaches to examine how the mouse homolog (mAMCase) can act in both acidic and neutral environments. We measured kinetic properties of mAMCase activity across a broad pH range, quantifying its unusual dual activity optima at pH 2 and 7. We also solved high resolution crystal structures of mAMCase in complex with oligomeric GlcNAcn, the building block of chitin, where we identified extensive conformational ligand heterogeneity. Leveraging these data, we conducted molecular dynamics simulations that suggest how a key catalytic residue could be protonated via distinct mechanisms in each of the two environmental pH ranges. These results integrate structural, biochemical, and computational approaches to deliver a more complete understanding of the catalytic mechanism governing mAMCase activity at different pH. Engineering proteins with tunable pH optima may provide new opportunities to develop improved enzyme variants, including AMCase, for therapeutic purposes in chitin degradation.

biochemistry↗