New carbohydrate binding domains identified by phage display based functional metagenomic screens of human gut microbiota

• Published in: Communications biology Vol. 6; no. 1; pp. 371 - 15
• Main Authors: Akhtar, Akil, Lata, Madhu, Sunsunwal, Sonali, Yadav, Amit, LNU, Kajal, Subramanian, Srikrishna, Ramya, T. N. C.
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 05.04.2023; Nature Publishing Group; Nature Portfolio
• Subjects: 45/22; 45/23; 45/29; 45/47; 45/77; 631/1647/2163; 631/326/2565/2142; 631/45/221; 631/45/612/1235; 631/45/72/1201; 82/16; 82/29; 82/47; 82/58; 82/80; 82/81; 82/83; Bacteriophages; Biology; Biomedical and Life Sciences; Carbohydrates; Commensals; Feces; Gastrointestinal Microbiome - genetics; Genes; Genomics; Glycoconjugates; Homology; Humans; Intestinal microflora; Levan; Life Sciences; Metagenomics; Microbiota; Phage display; Polysaccharides; Proteins
• ISSN: 2399-3642; 2399-3642
• DOI: 10.1038/s42003-023-04718-0

Uncultured microbes represent a huge untapped biological resource of novel genes and gene products. Although recent genomic and metagenomic sequencing efforts have led to the identification of numerous genes that are homologous to existing annotated genes, there remains, yet, an enormous pool of unannotated genes that do not find significant sequence homology to existing annotated genes. Functional metagenomics offers a way to identify and annotate novel gene products. Here, we use functional metagenomics to mine novel carbohydrate binding domains that might aid human gut commensals in adherence, gut colonization, and metabolism of complex carbohydrates. We report the construction and functional screening of a metagenomic phage display library from healthy human fecal samples against dietary, microbial and host polysaccharides/glycoconjugates. We identify several protein sequences that do not find a hit to any known protein domain but are predicted to contain carbohydrate binding module-like folds. We heterologously express, purify and biochemically characterize some of these protein domains and demonstrate their carbohydrate-binding function. Our study reveals several previously unannotated carbohydrate-binding domains, including a levan binding domain and four complex N-glycan binding domains that might be useful for the labeling, visualization, and isolation of these glycans. A metagenomic phage display library from healthy human fecal samples with carbohydrate binding screens identifies protein sequences of novel carbohydrate binding domains and their folds with AlphaFold2 predictions.