Microorganisms underpin modern industrial biotechnology; however, wild-type strains frequently exhibit suboptimal metabolic flux distribution, regulatory constraints, and limited product titers. To address these limitations, strain improvement strategies have evolved from classical mutagenesis and recombination approaches to precision genome-scale engineering. Recombinant DNA methodologies, including protoplast fusion, transduction, and conjugation, combined with advanced CRISPR-based systems (Cas9, Cas12, base editing, and prime editing), now enable programmable, locus-specific genomic modification with high fidelity. The integration of multi-omics platforms (genomics, epigenomics, transcriptomics, proteomics, metabolomics, and regulomics) with single-cell sequencing, fluxomics, and constraint-based metabolic modeling has transformed strain engineering into a predictive and systems-driven discipline. These approaches facilitate the rational rewiring of metabolic pathways, flux balancing, and regulatory circuits to enhance productivity and robustness. This review examines the progression from conventional strain improvement to genome-scale design strategies and evaluates methodologies for optimizing bacterial and fungal cell factories. It further discusses industrial translation, scalability constraints, and emerging opportunities for developing resilient microbial platforms for sustainable and carbon-efficient biomanufacturing.
Brewer’s spent grain (BSG), the major solid by-product of the brewing industry, is a nutrient-rich lignocellulosic biomass whose efficient and reproducible valorisation is challenged by its recalcitrant structure and pronounced compositional heterogeneity and batch-to-batch variability, arising from differences in cereal type and cultivar, type and proportion of malts and adjuncts used, beer style, and malting and mashing conditions. Unlike previous BSG-based fungal solid-state fermentation (SSF) studies that have generally focused on either enzyme production or compositional modification of the fermented solid, this study integrates xylanase production, partial purification and hydrolytic evaluation with characterisation of the resulting fermented BSG within a single SSF-based valorisation framework. Xylanase activity reached a maximum of (16.47 ± 1.04) kU g⁻¹ dry matter (DM) after 10 days of SSF. The crude enzyme extract was freeze-dried and subsequently partially purified by ammonium sulphate precipitation followed by dialysis, resulting in a 2.91-fold increase in specific activity and an activity recovery of 39.48
α-Arbutin is a skin-whitening compound widely used in cosmetic and skincare products due to its superior tyrosinase inhibitory activity and improved safety profile compared with hydroquinone. Although the use of amylosucrase for the enzymatic glycosylation of hydroquinone into α-arbutin represents an effective approach for α-arbutin production, the process remains limited by hydroquinone oxidation, low catalytic efficiency, and the requirement for antioxidant supplementation. In this study, a structure-guided protein engineering strategy was employed to improve the performance of amylosucrase from Chloroflexus islandicus (CiAS) for α-arbutin biosynthesis. Eleven CiAS variants were designed based on oxidation-prone surface residues and computational stability predictions and subsequently expressed in Escherichia coli. Among the engineered variants, CiAS-M250A exhibited the highest catalytic performance under the tested whole-cell biocatalytic conditions, reaching 250.2
The growing concern of antimicrobial-resistant (AMR) bacteria has created an urgent need for new antimicrobial agents with novel mechanisms of action. Non-ribosomal peptides (NRPs) have gained significant attention because they target essential and highly conserved bacterial structures through diverse mechanisms of action against many multidrug-resistant (MDR) pathogens. NRPs have distinct and diverse modes of action that reduce the likelihood of rapid AMR among bacteria. Advances in multi-omics approaches have greatly improved the discovery of novel NRPs In addition, synthetic biology, NRPS and domain engineering, and heterologous expression systems have expanded the possibilities for producing new peptide analogs with improved biological properties. This review discusses the major microbial sources of NRPs, their biosynthesis, recent approaches in NRP discovery and engineering, and their mechanisms of action against MDR pathogens along with challenges associated with NRP development, including silent biosynthetic gene clusters, low production yield, complex biosynthetic pathways, toxicity, and large-scale production with future prospects for developing NRP-based therapeutics to combat antimicrobial resistance.
Programmed cell autolysis can simplify the recovery of intracellular recombinant proteins, but differences in experimental conditions and evaluation criteria often limit direct comparisons of lysis systems. This study presents a multicriteria comparison of the two distinct lysis genes, the phage lambda SRRz cassette and the engineered 2PaT4L chimeolysin under identical IPTG-inducible conditions in Escherichia coli. The 2PaT4L outperformed the SRRz across multiple metrics, including cell viability, turbidity, protein release, and escape frequency. Using the inverse of escape frequency as a quantitative measure of lysis efficiency, 2PaT4L and SRRz reached (99.99 ± 0.012)
