Biology subjects

Gin, J.

Publications and source records attributed to Gin, J..

3 recordsLinked to original sources

bioRxiv2026

Generative Design of New-to-nature Biosynthetic Assembly Lines with Genomic Language Modeling

Reprogramming biosynthetic assembly lines can extend biosynthesis beyond the chemical space explored by nature. However, this remains difficult because assembly-line function depends on coordinated interactions across large multidomain enzymes. Here, we couple gLM2, a genomic language model trained on metagenomic sequences, with discrete diffusion and domain-level conditioning to enable generative design and optimization of biosynthetic gene clusters. We apply this approach to a chimeric type I polyketide synthase (PKS) engineered to produce {delta}-valerolactam, a molecule not naturally synthesized by PKSs. Through iterative redesign of two multi-domain regions in the context of the full PKS sequence, gLM2 progressively improved {delta}-valerolactam production, yielding variants with up to 9.4-fold higher titer than the starting enzyme. Together, these results demonstrate that evolutionary sequence information can be learned and applied to complex, multi-domain enzyme design problems, expanding biosynthetic assembly lines to produce molecules outside their natural biosynthetic repertoire.

bioengineering↗

bioRxiv2026

Expanding the scope of redox-balance growth coupling techniques with a carbon cofeeding strategy

Metabolic engineering to produce molecules not naturally synthesized by the host often requires directed evolution to improve pathway enzyme performance. Growth-coupled selection can dramatically increase directed-evolution throughput, and manipulation of redox balance has proven effective for tying reductase fitness to microbial growth. However, most redox-balance selections require feeding the reductase substrate because of stoichiometric constraints. This is impractical for many biosynthetic pathways either due to practical limitations on cost or complexity of bulk substrate synthesis, or the lack of an ability to transport substrate into cells, for example intracellular acyl-CoA/ACP intermediates. Here we define stoichiometric constraints that make substrate feeding necessary for many acetyl-CoA-derived reduction pathways in NADPH-imbalanced hosts. We overcome these constraints with a dual-feedstock strategy in which glucose provides reducing power while acetate supplies additional acetyl-CoA without directly perturbing redox balance. In an engineered E. coli selection strain, acetate co-feeding enabled growth coupling of acetaldehyde, 3-hydroxybutyrate, and mevalonate production and produced a linear correlation between product formation and growth. We then used this selection to evolve a class II HMG-CoA reductase (HMGR) from Delftia acidovorans toward NADPH utilization, enriching variants with improved NADPH-dependent activity. Finally, propionate co-feeding enabled growth coupling of propionyl-CoA reduction, supporting the generality of carbon co-feeding for selecting enzymes in pathways involving acyl-chain elongation and reduction. HighlightsO_LIStoichiometric limits of redox-balance growth coupling are defined C_LIO_LIAcetate co-feeding supplies acetyl-CoA without perturbing redox balance C_LIO_LICo-feeding enables growth coupling of acetaldehyde, 3-HB, and mevalonate C_LIO_LIGrowth coupling enables evolution of HMGR toward NADPH specificity C_LIO_LIPropionate co-feeding extends growth coupling to additional acyl-CoA substrates C_LI

bioengineering↗

bioRxiv2023

Ensemble and Iterative Engineering for Maximized Bioconversion to the Blue Pigment, Indigoidine from Non-Canonical Sustainable Carbon Sources

While many heterologous molecules can be produced via microbial bioconversion processes, maximizing their titers, rates, and yields from lignin-derived carbon streams remains challenging. Growth coupling can not only increase titers and yields but also shift the production period from stationary phase to growth phase. These methods for designing growth-coupling strains however require multi-gene edits for implementation which may be perceived as impractical. Here, we computationally evaluated 4,114 potential solutions for growth coupling para-coumarate to indigoidine production and prototype two cut sets in Pseudomonas putida KT2440. We used adaptive laboratory evolution (ALE) on the initial triple deletion strain to restore growth on p-CA. Using X-ray tomography on this post-ALE strain we revealed increased cell density and decreased cell volume. Proteomics identified upregulated peroxidases that mitigate reactive oxygen species formation. Nine iterative stepwise modifications further informed by model-guided and rational approaches realized a growth coupled strain that produced 7.3 g/L indigoidine at 77% MTY in para-coumarate minimal medium. These ensemble strategies provide a blueprint for producing target molecules at high product titers, rates, and yields. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=80 SRC="FIGDIR/small/532821v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@f0754aorg.highwire.dtl.DTLVardef@a45d24org.highwire.dtl.DTLVardef@df0596org.highwire.dtl.DTLVardef@1e7e140_HPS_FORMAT_FIGEXP M_FIG C_FIG

systems biology↗