# Engineering Methylotrophic Yeast for in vivo Production of Deoxysugars from DHAP

> **NIH NIH R43** · MIDWEST BIOPROCESSING CENTER, LLC · 2020 · $193,138

## Abstract

The goal of this proposal is to develop a fermentation system to produce a variety of rare and
deoxysugars using promiscuous DHAP-dependent aldolases. Carbon-carbon bond forming aldolase reactions
are of great relevance to the preparation of fine chemicals, intermediates and bioactive chemicals. The
aldolase enzymes responsible for this kind of C-C bond formation catalyze an asymmetric aldol condensation
which consists of coupling an aldehyde and a ketone. Dihydroxyacetone phosphate (DHAP) dependent
aldolases are attractive because there are four different types available with different stereo-specificities. While
substrate specificity for the acceptor is flexible, they require DHAP to be the donor. Unfortunately, DHAP is
very unstable and expensive, making it impractical for industrial use.
 We propose to develop a novel approach to produce DHAP using a metabolically modified strain of
Hansenula polymorpha, a methylotrophic yeast. H. polymorpha has a unique metabolism with the ability create
C6 and DHAP solely from mixed carbon sources including methanol. Mutants that that accumulate the
precursor to DHAP, dihydroxyacetone at high yields have been identified and will be used to enhance DHAP
production.
 In Phase I we will demonstrate the feasibility of using an in vivo conversion of intracellular DHAP to
produce the model compound L-fuculose. This reaction is catalyzed by Fuculose-1-phosphate aldolase (FucA).
Specifically, we will demonstrate the ability to produce L-fuculose using H. polymorpha cell-free extracts,
engineer H. polymorpha for improved DHAP production, clone and express the fuculose-1-phosphate aldolase
into H. polymorpha, and finally demonstrate the production of L-fuculose in vivo.
 In Phase II we will optimize the system further, knock-out genes that convert DHAP to GAP, and
introduce fuculose isomerase for production of L-fucose. We will then clone the other main DHAP-dependent
aldolases with broader specificity to aldehydes and demonstrate production of dozens of deoxysugars, rare
sugars and carbohydrates using different aldehyde acceptors. In Phase III the technology will be
commercialized by licensing DHAP and aldolase producing strains and carrying out contract R&D using the
system for the generation of drug discovery candidates and the preparative of pharmaceutical intermediates.

## Key facts

- **NIH application ID:** 10082147
- **Project number:** 1R43GM139456-01
- **Recipient organization:** MIDWEST BIOPROCESSING CENTER, LLC
- **Principal Investigator:** Leila Aminova
- **Activity code:** R43 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $193,138
- **Award type:** 1
- **Project period:** 2020-09-01 → 2021-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10082147

## Citation

> US National Institutes of Health, RePORTER application 10082147, Engineering Methylotrophic Yeast for in vivo Production of Deoxysugars from DHAP (1R43GM139456-01). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10082147. Licensed CC0.

---

*[NIH grants dataset](/datasets/nih-grants) · CC0 1.0*