Activity-Based Tracking of Glycan Turnover in Microbiomes

Area of Science:

• Microbiology
• Biochemistry
• Environmental Science

Background:

• Glycans are crucial components of microbial ecosystems, influencing the global carbon cycle and human health.
• Current methods for tracking microbial glycan turnover and identifying active degraders are limited, especially without prior genomic information.

Purpose of the Study:

• To develop and validate a novel activity-based fluorescence resonance energy transfer (FRET) probe for direct visualization and quantification of glycan metabolism in complex microbial communities.
• To demonstrate the probe's utility in tracking α-mannan degradation, a key polysaccharide in algal blooms.

Main Methods:

• Automated glycan assembly was used to synthesize a mannan hexasaccharide functionalized with a fluorescein-rhodamine FRET pair.
• The FRET probe was validated using a recombinant endo-α-mannanase (GH76) in cell lysates, pure cultures, and complex microbiomes.
• Microscopy and plate assays were employed to visualize and quantify in situ glycan turnover.

Main Results:

• The FRET probe successfully enabled visualization and quantification of α-mannan degradation.
• The probe demonstrated functionality across various biological contexts, from purified enzymes to complex marine microbiomes.
• Spatiotemporal dynamics of α-mannan turnover were visualized in a marine microbial community.

Conclusions:

• The developed glycan FRET probes are versatile tools for studying glycan metabolism.
• These probes facilitate tracking of polysaccharide degradation across diverse biological scales, from enzymes to entire microbiomes.
• This technology offers new possibilities for understanding microbial roles in biogeochemical cycles and host-microbe interactions.