Economical Modular One-Pot Multienzyme Synthesis of Human Milk Oligosaccharides

经济的模块化一锅多酶合成母乳低聚糖

基本信息

  • 批准号:
    10575228
  • 负责人:
  • 金额:
    $ 22.65万
  • 依托单位:
  • 依托单位国家:
    美国
  • 项目类别:
  • 财政年份:
    2023
  • 资助国家:
    美国
  • 起止时间:
    2023-05-01 至 2025-04-30
  • 项目状态:
    未结题

项目摘要

Project Summary Glycans play essential roles in virtually all aspects of biology. Thus, they have broad applications, including supplementation of necessary human milk oligosaccharides (HMOs) to infant formula as therapeutics to prevent infection by multiple pathogens, maintain proper microbiota, prevent allergies, and treat necrotizing enterocolitis in infants. However, present synthesis methods for the more complex HMOs fail in affordable scalability that hinders their development as glycan therapeutics. Although current one-pot multienzyme (OPME) systems for HMO synthesis have streamlined numerous synthetic reaction steps, little has been done on optimization and scalability. We hypothesize that a modular OPME system for glycan synthesis can be optimized and integrated with inexpensive polyphosphate-based energy regeneration to decrease cost and significantly increase yield of desirable glycans. In Aim 1, we will establish reference OPME reactions for single sugar transfer steps that start from simple building blocks of monosaccharides and glycan acceptors as input, and provide enzymes for high- energy sugar donor synthesis, monosaccharide transfer and energy regeneration. These studies will develop scalable modules for optimization of chemical input (donor and acceptor building blocks, catalytic quantities of nucleotides, divalent cations, controlled pH, and enzyme catalysts) to establish cross-platform compatible reaction conditions. In Aim 2, we will establish a scalable, coupled polyphosphate energy regeneration system for OPME glycan synthesis. Optimization of polyphosphate as an energy source will require control of polyphosphate and divalent cation concentrations, neutralization of pH changes, and integration of enzymes that generate ATP (RpPPK2-3) and distribute high-energy phosphate equivalents to other nucleotide forms (NDK). Our goals are to create a universal energy source that integrates the continual synthesis of UDP-, GDP-, CMP-, and ADP-sugar donors in large-scale OPME reactions. In Aim 3, we will generate proof-of-concept scalable OPME synthesis of model HMO targets of biological interest for optimization. The approach will combine synthetic modules for sugar donor synthesis and glycan extension with optimized energy regeneration and determine conditions for cross-platform compatibility for all enzymatic steps in the energy-coupled OPME (ecOPME) platform. The HMO targets also provide opportunities to test combined ecOPME reactions as well as sequential ecOPME sugar additions where enzyme competition would yield undesired products. The goals are to integrate multiple enzymatic transfer steps through the selective use of glycosyltransferase acceptor specificity coupled with energy regeneration to result in a proof-of-concept modular platform for efficient, flexible, and scalable synthesis of target therapeutic HMOs starting from simple monosaccharide building blocks.
项目总结

项目成果

期刊论文数量(0)
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科研奖励数量(0)
会议论文数量(0)
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KELLEY W. MOREMEN其他文献

KELLEY W. MOREMEN的其他文献

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{{ truncateString('KELLEY W. MOREMEN', 18)}}的其他基金

2013/2015 Glycobiology Gordon Research Conference & Gordon Research Seminar
2013/2015 糖生物学戈登研究会议
  • 批准号:
    8451685
  • 财政年份:
    2013
  • 资助金额:
    $ 22.65万
  • 项目类别:
QRT-PCR TRANSCRIPT ANALYSIS
QRT-PCR 转录本分析
  • 批准号:
    8363006
  • 财政年份:
    2011
  • 资助金额:
    $ 22.65万
  • 项目类别:
REGULATION OF ERAD- AND UPR-RELATED GENE EXPRESSION
ERA 和 UPR 相关基因表达的调控
  • 批准号:
    8363017
  • 财政年份:
    2011
  • 资助金额:
    $ 22.65万
  • 项目类别:
MICROARRAY VALIDATION OF DATA FROM WILD-TYPE AND MGAT5 KNOCK-OUT MOUSE TISSUES
野生型和 MGAT5 敲除小鼠组织数据的微阵列验证
  • 批准号:
    8363111
  • 财政年份:
    2011
  • 资助金额:
    $ 22.65万
  • 项目类别:
EXPRESSION/LABELING OF GLYCOPROTEINS FOR NMR-BASED STRUCTURE STUDIES
用于基于 NMR 的结构研究的糖蛋白表达/标记
  • 批准号:
    8361783
  • 财政年份:
    2011
  • 资助金额:
    $ 22.65万
  • 项目类别:
N-GLYCAN PROCESSING ENZYMES IN GLYCOPROTEIN MATURATION & QUALITY CONTROL
糖蛋白成熟中的 N-聚糖加工酶
  • 批准号:
    8361789
  • 财政年份:
    2011
  • 资助金额:
    $ 22.65万
  • 项目类别:
REGULATION OF TRANSCRIPTS RELATED TO RAT ASN-LINKED GLYCAN BIOSYNTHESIS
与大鼠 ASN 连接聚糖生物合成相关的转录调控
  • 批准号:
    8363015
  • 财政年份:
    2011
  • 资助金额:
    $ 22.65万
  • 项目类别:
ANALYSIS OF TRANSCRIPTS INVOLVED IN GLYCOCONJUGATE SYNTHESIS IN D MELANOGASTER
黑腹果蝇糖复合物合成中涉及的转录本分析
  • 批准号:
    8363041
  • 财政年份:
    2011
  • 资助金额:
    $ 22.65万
  • 项目类别:
CHANGES IN GENE EXPRESSION IN NDST1 & NDST2 KNOCK-OUT MOUSE CELLS
NDST1 中基因表达的变化
  • 批准号:
    8363110
  • 财政年份:
    2011
  • 资助金额:
    $ 22.65万
  • 项目类别:
BIOMARKER DISCOVERY FOR PATHOLOGICAL PLACENTAL MALARIA
病理性胎盘疟疾生物标志物的发现
  • 批准号:
    8363040
  • 财政年份:
    2011
  • 资助金额:
    $ 22.65万
  • 项目类别:
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