Abstract:

Human milk oligosaccharides (HMOs), recognized as key bioactive constituents of human milk, play indispensable roles in infant growth and development through immunomodulation, gut microbiota regulation, and pathogen defense mechanisms. These multifaceted compounds have emerged as pivotal ingredients in next-generation infant formula formulations. Current investigations primarily concentrate on three domains: elucidation of biological functions, exploration of therapeutic applications, and optimization of biosynthesis platforms. The growing commercial demand for HMOs has driven significant advancements in enzymatic synthesis and metabolic engineering approaches to achieve cost-effective production at industrial scales. The rapid development of synthetic biology has revolutionized the production of HMOs, making large-scale microbial fermentation a viable and economically feasible strategy for industrial applications. This paradigm shift has significantly accelerated the commercialization of HMOs, which are increasingly recognized for their critical roles in infant nutrition, gut microbiota modulation, and immune system development. In this comprehensive review, we systematically evaluate the latest research progress in HMOs, with particular emphasis on their biosynthesis, functional mechanisms, and biotechnological production, focusing on four key dimensions: (1) Physiological functions and mechanistic insights of HMOs-recent studies have elucidated the prebiotic, antimicrobial, and immunomodulatory properties of HMOs, highlighting their structural diversity and structure-function relationships. Advances in glycomics and microbiome research have deepened our understanding of how HMOs influence host-microbe interactions and metabolic pathways. (2) Discovery and engineering of glycosyltransferases for HMOs biosynthesis-glycosyltransferases (GTs) play a pivotal role in determining the structural diversity of HMOs. We review the latest strategies for enzyme mining, characterization, and protein engineering, including directed evolution and computational design, to enhance catalytic efficiency and substrate specificity. (3) Metabolic pathway design and optimization in microbial cell factories–the construction of efficient microbial chassis for HMOs production requires systematic pathway engineering. Key considerations include precursor supply, cofactor balancing, and the elimination of metabolic bottlenecks. We discuss recent advances in dynamic pathway regulation and modular assembly techniques to improve yield and purity. (4) Synthetic biology strategies for chassis cell development–the optimization of host strains involves genome-scale metabolic modeling, CRISPR-based genome editing, and adaptive laboratory evolution. Additionally, novel approaches such as cell-free biosynthesis and consortia-based fermentation are emerging as promising alternatives for complex HMOs synthesis. Furthermore, we discuss emerging challenges and future directions in this field.

Key words: human milk oligosaccharides, glycosyltransferases, synthetic biology, metabolic network regulation

摘要：

母乳低聚糖（Human milk oligosaccharides，HMOs）是母乳中的核心营养成分，对于婴幼儿的生长发育具有无可替代的生理功效，是新一代婴幼儿配发奶粉的核心原料。目前针对HMOs的研究主要集中在生理功能、临床应用以及生物合成技术开发等方面。鉴于HMOs广阔的市场需求，高效生物合成HMOs逐渐成为研究热点。合成生物学技术的突破使微生物发酵法大规模生产HMOs成为可能，显著提升了产业化进程的经济可行性。本文系统综述近年来HMOs的研究进展并展望了HMOs的发展趋势及挑战，包括：（1）HMOs相关生理功能研究进展；（2）HMOs生物合成关键糖基转移酶研究进展；（3）基于微生物细胞工厂的HMOs生物合成路径设计；（4）用于改造HMOs合成底盘细胞的合成生物学策略。通过解析HMOs生物合成关键酶元件筛选、代谢途径通量平衡及底盘细胞代谢网络调控等核心科学问题，为HMOs的高效生物制造提供理论依据与技术参考。

关键词: 母乳低聚糖, 糖基转移酶, 合成生物学, 代谢网络调控