7qee: Difference between revisions
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The abundance of recorded protein sequence data stands in contrast to the small number of experimentally verified functional annotation. Here we screened a million-membered metagenomic library at ultrahigh throughput in microfluidic droplets for beta-glucuronidase activity. We identified SN243, a genuine beta-glucuronidase with little homology to previously studied enzymes of this type, as a glycoside hydrolase 3 family member. This glycoside hydrolase family contains only one recently added beta-glucuronidase, showing that a functional metagenomic approach can shed light on assignments that are currently 'unpredictable' by bioinformatics. Kinetic analyses of SN243 characterized it as a promiscuous catalyst and structural analysis suggests regions of divergence from homologous glycoside hydrolase 3 members creating a wide-open active site. With a screening throughput of >10(7) library members per day, picolitre-volume microfluidic droplets enable functional assignments that complement current enzyme database dictionaries and provide bridgeheads for the annotation of unexplored sequence space. | The abundance of recorded protein sequence data stands in contrast to the small number of experimentally verified functional annotation. Here we screened a million-membered metagenomic library at ultrahigh throughput in microfluidic droplets for beta-glucuronidase activity. We identified SN243, a genuine beta-glucuronidase with little homology to previously studied enzymes of this type, as a glycoside hydrolase 3 family member. This glycoside hydrolase family contains only one recently added beta-glucuronidase, showing that a functional metagenomic approach can shed light on assignments that are currently 'unpredictable' by bioinformatics. Kinetic analyses of SN243 characterized it as a promiscuous catalyst and structural analysis suggests regions of divergence from homologous glycoside hydrolase 3 members creating a wide-open active site. With a screening throughput of >10(7) library members per day, picolitre-volume microfluidic droplets enable functional assignments that complement current enzyme database dictionaries and provide bridgeheads for the annotation of unexplored sequence space. | ||
Functional metagenomic screening identifies an unexpected beta-glucuronidase.,Neun S, Brear P, Campbell E, Tryfona T, El Omari K, Wagner A, Dupree P, Hyvonen M, Hollfelder F Nat Chem Biol. 2022 Oct;18(10):1096-1103. doi: 10.1038/s41589-022-01071-x. Epub, 2022 Jul 7. PMID:35799064<ref>PMID:35799064</ref> | Functional metagenomic screening identifies an unexpected beta-glucuronidase.,Neun S, Brear P, Campbell E, Tryfona T, El Omari K, Wagner A, Dupree P, Hyvonen M, Hollfelder F Nat Chem Biol. 2022 Oct;18(10):1096-1103. doi: 10.1038/s41589-022-01071-x. Epub , 2022 Jul 7. PMID:35799064<ref>PMID:35799064</ref> | ||
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | ||
Latest revision as of 06:42, 21 November 2024
SN243 mutant D415N bound to para-nitrophenyl-Beta-D-glucuronide
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