Critical Reviews in Biotechnology最新文献

Lytic polysaccharide monooxygenases (LPMOs) are auxiliary metalloenzymes that play a crucial role in the degradation of polysaccharides through an oxidative mechanism, distinguishing them from the traditional glycoside hydrolases. Although LPMOs were first identified in 1992, their functional identity and unique oxidative activity were not fully understood until the year 2010. These enzymes cleave at the C1 or C4 position of glycosidic bonds in polysaccharides using molecular oxygen and reductants such as ascorbic acid or cellobiose dehydrogenase (CDH). LPMOs exhibit significant sequence diversity across eight known families and operate via complex mechanisms. Structurally, LPMOs have a conserved active site with a copper ion coordinated by two histidine residues, known as the "histidine brace" which is crucial for their oxidative activity Their ability to enhance the efficiency of cellulase enzymes makes them highly valuable in the bio refinery industry. This review focuses on details of: regioselectivity, reaction mechanism, protein engineering strategies, and industrial applications of LPMO. It also emphasizes the building correlation between challenges at the industrial level and their possible solutions through enzyme engineering.

Enzymes are highly efficient biocatalysts known for their chemical, regio-, and stereoselectivity, making them valuable in industrial applications. While directed evolution has expanded the scope of enzyme-catalyzed reactions, the range of enzymatic reactions remains limited compared to reactions catalyzed by chemical catalysts. Computational enzyme design has achieved de novo enzyme design, but the approach is often complex, time-intensive, and has a low success rate. A promising strategy to design novel enzymes involves developing noncanonical amino acids (ncAAs) with catalytic potential and integrating them into protein scaffolds via genetic codon expansion technology. This method combines the novel reactivity of ncAAs with the high selectivity provided by protein scaffolds, significantly enhancing the diversity of enzyme-catalyzed reactions. This review discusses recent advancements in novel enzyme design using ncAAs, including those being used as catalytic groups, metal-coordinating groups, heme ligands, and photocatalytic groups. The article emphasizes the broad potential of using ncAAs in enzyme design to expand the diversity of enzyme-catalyzed reactions, and outlooks the potential applications of artificial intelligence technology in this area.

Polymerase chain reaction (PCR) is a critical technology in nucleic acid detection and quantification. The PCR reaction requires thermal cycling the reaction mixture between two or more temperature stages for ∼30 cycles to achieve exponential amplification of the target DNA. Typically, the thermal cycling takes roughly an hour to finish and the large time consumption is a drawback for PCR. We review the various methods developed to reduce the thermal cycling time and build a rapid PCR. We group the methods to two approaches. The first approach is to increase the local heating/cooling power. The methods in this approach include contact heating, such as: heating resistors and Peltier pumps, and non-contact heating using air-blow, radiation on water and plasmonics. The other approach is to rapidly move the reaction mixture to a different temperature zone. Methods in this approach include: relocating the reaction vessel, continuous flow PCR using microfluidic chips, long tubes or oscillatory PCR scheme, and convective PCR. We analyze the advantages and challenges for each method used and the critical parameters to consider when evaluating the technologies. We review the technological advances and commercialization for each method. We also discuss the current challenges and future directions in building an effective and commercial rapid PCR, with the emphasis on sensitivity, portability and cost.

The escalating problem of antibiotic resistance has sparked renewed interest in bacteriophages (phages) as potential substitutes for conventional antibiotics in treating infectious diseases, improving food safety, and advancing sustainable agriculture. The key phage research processes, such as host range, burst size, and environmental stability tests, strongly influence phage production processes. Hence, the standardization of the mentioned techniques must be prioritized. The introduction of high-throughput sequencing technologies with high accuracy and the emergence of novel bioinformatic tools to analyze the resulting raw molecular data provide comprehensive identification of phages and phage-verse (the universe of phage). While encapsulation of phages was studied comprehensively before, the production of encapsulated phages is still unclear. Moreover, recent advances in artificial intelligence (AI) contribute to phage research by increasing the accuracy of bioinformatic tools, improving resistance profiling, and facilitating phage host prediction. Incorporating AI promises a future of automated, precisely tailored phage applications. This review covers efficient techniques appropriate for industrial and agricultural applications as well as large-scale phage production methods, covering upstream and downstream processing. Also, encapsulated phage production and AI-based automated systems in various applications are proposed in this review. By covering both present issues and potential future uses of phages in the fight against antibiotic resistance, this review seeks to give academics and industry experts the fundamental information they need to advance phage-based solutions.

In recent years, interest in the role of nutrient cycling in sustainable agriculture has significantly increased. The potential of arbuscular mycorrhizal (AM) fungi (AMFs) in nutrient cycling and plant growth improvement has long been recognized. However, there have been only a few studies on the identification and exploration of AM symbiosis-related plant genes for sustainable agriculture. We have developed a new constructive model for using host plant-derived AM symbiosis-related genes to improve breeding and AMF utilization for sustainable agriculture, particularly in the context of climate change. This model include: 1) the discovery of AM symbiosis-related genes in crop wild-relatives for molecular breeding and 2) the screening and propagation of AMFs that can help improve water-use efficiency and nutrient-use efficiency by crops, thereby reducing chemical fertilizer use in agricultural production. The first approach uniquely facilitates the identification of host plant-derived AM symbiosis-related genes, such as CHITIN ELICITOR RECEPTOR KINASE 1 (OsCERK1) from Dongxiang (DY) wild rice (Oryza rufipogon) (OsCERK1DY), MILDEW RESISTANCE LOCUS 1 (MLO1) from wild barley (Hordeum spontaneum), and WRKY60 from wild soybean (Glycine soja), for breeding purposes. The second one involves identifying soil-borne AMF species, such as Rhizophagus intraradices and Glomus mosseae for practical applications in the field. This suggestive model presents an emerging biotechnological potential for engineering climate-resilient crops.

Astaxanthin, a natural di-keto carotenoid xanthophyll, is a highly valued nutraceutical and food ingredient due to its potent health benefits, including: anti-inflammatory, antioxidant, anti-cancer, cardiovascular, and anti-diabetic effects. This review examines the primary natural sources of: astaxanthin microalgae, yeast, bacteria, and plants, with a focus on microalgae due to their superior accumulation potential and bioactivity. It explores the growing prospects for large-scale astaxanthin production, highlighting advancements in both upstream and downstream processes. Upstream innovations include enhanced bioprocess designs that improve biomass yield, light and stress tolerance. Downstream, sustainable extraction methods such as aqueous two-phase systems with deep eutectic solvents (99.64% recovery) and high-pressure supercritical CO₂ extraction have improved efficiency and scalability. Additionally, eco-friendly techniques, such as bead milling and pulsed electric field permeabilization offer cost-effective solutions, among other cell disruption techniques, and ensure higher yields. This study provides a comprehensive overview of recent advances in astaxanthin production and extraction, aligned with the Sustainable Development Goals (SDGs) linked to health and well-being (SDG 3) and responsible consumption and production (SDG 12).

Despite their tremendous benefits to society, currently licensed vaccines, including mRNA-based ones, are far from ideal and suffer several issues. Common problems associated with all types of vaccine formulations currently in clinical use include thermolabile nature, poor shelf life at ambient temperature, and the continuous need for cold chain and sometimes ultra-low temperature. Several approaches have been tested in the past to surmount these shortcomings. This review discusses the advantages of whole yeast (WY) or whole recombinant yeast-based (WRY) vaccines compared to other vaccine formulations to overcome the above-mentioned issues. The interaction between yeast cells and the host immune system in relevance to the WRY vaccines has been discussed along with the importance of whole yeast cells in the development of anti-fungal vaccines by highlighting the bottlenecks hampering the use of WRY in vaccine formulation. Specifically, the present review highlighted the status of WRY vaccines, including those in clinical trials, and also summarized the guidelines, one should follow while conducting research or reporting the data related to WRY vaccines.

Reactive oxygen species (ROS) play crucial roles in many plant biological processes. ROS have emerged as major signaling molecules mediating various regulatory reactions in response to environmental stimuli. This signaling is mediated by a highly regulated process of ROS accumulation at specific cellular compartments. Therefore, this review focuses on the intricate ROS signaling in plant defense and strategic virulence effectors from pathogens hijacking ROS homeostasis. In addition, the ROS-mediated regulation of plant processes acts through post-translational modifications (PTMs) is discussed. We also provide a valuable roadmap for translating ROS research into climate-resilient cultivars by exploring the manipulation of pathogen effectors, ROS cascade genes through modern biotechnological approaches, and post-translational modifications against various environmental stressors. This framework can advance research in plant stress biology and agricultural practices.

Urease (urea amidohydrolase, EC 3.5.1.5), first crystallized from jack-bean (Canavalia ensiformis) by James B. Sumner in 1926, has become a cornerstone of biotechnology. The global urease market, dominated by plant-based sources, was valued at USD 1.24 Billion in 2024 and is projected to grow at a CAGR of 5.5%, reaching USD 1.94 billion by 2033. However, plant-derived ureases face challenges, such as low extraction efficiency, variability in yield due to plant maturity, and sensitivity to environmental factors, limiting scalability. Microbial ureases, globally embraced due to escalating demand, offer superior stability across extreme pH and temperature ranges. These attributes confer broad potential applications in diverse fields, such as: agriculture, environmental, clinical, and healthcare industries. Nevertheless, the industrial production of microbial urease continues to encounter obstacles, including elevated purification costs and the lack of cost-effective optimization strategies. This review provides quantitative insights into microbial ureases from bacteria, fungi (excluding hemiascomyces), and diatoms, highlighting their catalytic efficiency, Ni-dependencies, and advancements in assay techniques and enhanced purification strategies. It explores applications across agriculture, bioremediation, and self-healing concrete, emphasizing ureolysis-driven microbially induced carbonate precipitation (MICP) as a promising eco-friendly and sustainable approach, thus providing a scientific and reasonable reference for their large-scale application.

Pellets are ultrastructural configurations of filamentous fungal biomass that form during growth in submerged culture. This growth pattern offers advantages for controlling and stabilizing bioprocesses through biomass immobilization, reduced medium viscosity, and facilitated compound extraction. These benefits are particularly valuable for bioremediation, synergistic applications with biomaterials, and industrial metabolite production. However, fungal pellets also present challenges, such as limited oxygen diffusion to the pellet core, inconsistent pellet homogeneity, and decreased productivity. Factors such as electrostatic interactions, hydrophobicity, and culture conditions influence pellet formation. Currently, optimization efforts for pellet production focus on evaluating parameters, such as: pH range, agitation rate, pellet formation time, carbon source, additive agents, trace metals, and inoculum concentration, among others. Fungal pellets are increasingly recognized as promising platforms in emerging environmental biotechnology due to their versatility in pollutant removal and sustainable processing. Unlike previous reviews, this work provides an integrated and up-to-date perspective that combines the fundamentals of pellet formation with recent advances in their environmental and industrial applications, highlighting strategies for optimization and scalability. This review focuses on analyzing the most relevant aspects of fungal pellets, including their formation, optimization, and biotechnological applications. Given the growing importance of fungi in contemporary biotechnology, a state-of-the-art review of fungal pellets is warranted. This includes presenting an updated overview of processes for generating fungal biomass with enhanced handling, based on the use of fungal granules, an essential component for the implementation of efficient biotechnological processes using model fungal pellets with relevant industrial applications.