Systems and methods for identifying and expressing gene clusters
Inventors
Naughton, Brian Thomas • Harvey, Colin • Schlecht, Ulrich • Hillenmeyer, Maureen Elizabeth • Horecka, Joe
Assignees
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Publication Number
US-12456540-B2
Publication Date
2025-10-28
Expiration Date
Abstract
Methods for identifying biosynthetic gene clusters that include genes for producing compounds that interact with specific target proteins are disclosed. Some methods relate to bioinformatics methods for identifying and/or prioritizing biosynthetic gene clusters. Related systems, components, and tools for the identification and expression of such gene clusters are also disclosed.
Core Innovation
The invention provides systems and methods for identifying fungal biosynthetic gene clusters that encode proteins similar to a first target protein. A fungal protein is identified based on its similarity to the first target protein, with at least 60% sequence identity, and a fungal gene cluster is identified that is within 20,000 bases of a genomic region that encodes the fungal protein.
The identified gene cluster is used to produce secondary metabolites and small molecules through heterologous expression in a host cell. The fungal gene cluster or a plurality of genes from the fungal gene cluster is expressed in a host cell to produce a compound, and the compound is isolated.
The isolated compound is screened for inhibition of an activity of the first target protein, wherein the first target protein is a mammalian protein. The document also describes cryptic fungal biosynthetic gene clusters, including type I PKS and UbiA-like UTC, heterologously expressed in a host cell, and reports computer-implemented BGC identification and ranking using homology and phylogenetic analysis alongside multiple scoring factors.
Claims Coverage
The claim coverage centers on four inventive features linked to identifying a fungal BGC associated with a mammalian target protein, producing a compound by host-cell expression, isolating the compound, and screening it for inhibition of target activity.
Target-protein similarity-guided fungal protein selection
Identifying a fungal protein based on its similarity to a first target protein, wherein the fungal protein has at least 60% sequence identity to the first target protein.
Genomic neighborhood BGC identification
Identifying a fungal gene cluster that is within 20,000 bases of a genomic region that encodes the fungal protein.
Heterologous BGC or gene-expression for compound production
Expressing the fungal gene cluster or a plurality of genes from the fungal gene cluster in a host cell to produce a compound.
Compound isolation and inhibition screening for a mammalian target
Isolating the compound produced by the fungal gene cluster or the plurality of genes from the fungal gene cluster; screening the isolated compound for inhibition of an activity of the first target protein, wherein the first target protein is a mammalian protein.
The claim coverage links a mammalian target protein to a fungal gene cluster by similarity and genomic proximity, expresses the gene cluster or selected genes in a host cell to produce a compound, isolates the compound, and screens it for inhibition of the target protein activity.
Stated Advantages
Measurable compounds are produced from productive clusters.
A productive cluster fraction of 54% is reported.
Non-native products are produced in S. cerevisiae.
Intron correction restores proper alignment and compound production in a host cell.
Documented Applications
Cryptic fungal biosynthetic gene clusters, including type I PKS and UbiA-like UTC, are heterologously expressed in a host cell.
BGCs are identified in fungal genomes and evaluated using untargeted metabolomics and compound structure determination approaches such as LC-MS and NMR.
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