Extracellular BSA-degrading SAPs in the rare pathogen Meyerozyma guilliermondii strain SO as potential virulence factors in candidiasis.

Meyerozyma guilliermondii (Candida guilliermondii) is one of the Candida species associated with invasive candidiasis. With the potential for expressing industrially important enzymes, M. guilliermondii strain SO possessed 99 % proteome similarity with the clinical ATCC 6260 isolate and showed pathogenicity towards zebrafish embryos. Recently, three secreted aspartyl proteinases (SAPs) were computationally identified as potential virulence factors in this strain without in vitro verification of SAP activity. The quantification of Candida SAPs activity in liquid broth were also scarcely reported. Thus, this study aimed to characterize M. guilliermondii strain SO's ability to produce SAPs (MgSAPs) in different conditions (morphology and medium) besides analyzing its growth profile. MgSAPs' capability to cleave bovine serum albumin (BSA) was also determined to propose that MgSAPs as the potential virulence factors compared to the avirulent Saccharomyces cerevisiae. M. guilliermondii strain SO produced more SAPs (higher activity) in yeast nitrogen base-BSA-dextrose broth compared to yeast extract-BSA-dextrose broth despite insignificantly different SAP activity in both planktonic and biofilm cells. FeCl3 supplementation significantly increased the specific protein activity (∼40 %). The BSA cleavage by MgSAPs at an acidic pH was proven through semi-quantitative SDS-PAGE, sharing similar profile with HIV-1 retropepsin. The presented work highlighted the MgSAPs on fungal cell wall and extracellular milieu during host infection could be corroborated to the quantitative production in different growth modes presented herein besides shedding lights on the potential usage of retropepsin's inhibitors in treating candidiasis. Molecular and expression analyses of MgSAPs and their deletion should be further explored to attribute their respective virulence effects.

Thermodynamics of a hyperthermostable carboxylesterase from Anoxybacillus geothermalis D9.

Hyperthermostable enzymes are highly desirable biocatalysts due to their exceptional stability at extreme temperatures. Recently, a hyperthermostable carboxylesterase EstD9 from Anoxybacillus geothermalis D9 was biochemically characterized. The enzyme exhibited remarkable stability at high temperature. In this study, we attempted to probe the conformational adaptability of EstD9 under extreme conditions via in silico approaches. Circular dichroism revealed that EstD9 generated new ß-sheets at 80 °C, making the core of the hydrolase fold more stable. Interestingly, the profiles of molecular dynamics simulation showed the lowest scores of radius of gyration and solvent accessible surface area (SASA) at 80 °C. Three loops were responsible for protecting the catalytic site, which resided at the interface between the large and cap domains. To further investigate the structural adaptation in extreme conditions, the intramolecular interactions of the native structure were investigated. EstD9 revealed 18 hydrogen bond networks, 7 salt bridges, and 9 hydrophobic clusters, which is higher than the previously reported thermostable Est30. Collectively, the analysis indicates that intramolecular interactions and structural dynamics play distinct roles in preserving the overall EstD9 structure at elevated temperatures. This work is relevant to both fundamental and applied research involving protein engineering of industrial thermostable enzymes.

Enhancement in T1 lipase purification recovery using the novel construct pGEX4T1/His-T1.

The Geobacillus zalihae strain T1 produces a thermostable T1 lipase that could be used for industrial purposes. Previously, the GST-T1 lipase was purified through two chromatographic steps: affinity and ion exchange (IEX) but the recovery yield was only 33%. To improve the recovery yield to over 80%, the GST tag from the pGEX system was replaced with a poly-histidine at the N-terminal of the T1 lipase sequence. The novel construct of pGEX/His-T1 lipase was developed by site-directed mutagenesis, where the XbaI restriction site was introduced upstream of the GST tag, allowing the removal of tag via double digestion using XbaI and EcoRI (existing cutting site in the pGEX system). Fragment of 6 × His-T1 lipase fusion was synthesized, cloned into the pGEX4T1 system, and expressed in Escherichia coli BL21 (DE3) pLysS, resulting in lipase-specific activity at 236 U/mg. The single purification step of His-T1 lipase was successfully achieved using nickel Sepharose 6FF with an optimized concentration of 5 mM imidazole for binding, yielding the recovery of 98%, 1,353 U/mg lipase activity, and a 5.7-fold increase in purification fold. His-T1 lipase was characterized and was found to be stable at pH 5-9, active at 70 °C, and optimal at pH 9.

Knotting terminal ends of mutant T1 lipase with disulfide bond improved structure rigidity and stability.

Lipase biocatalysts offer unique properties which are often impaired by low thermal and methanol stability. In this study, the rational design was employed to engineer a disulfide bond in the protein structure of Geobacillus zalihae T1 lipase in order to improve its stability. The selection of targeted disulfide bond sites was based on analysis of protein spatial configuration and change of Gibbs free energy. Two mutation points (S2C and A384C) were generated to rigidify the N-terminal and C-terminal regions of T1 lipase. The results showed the mutant 2DC lipase improved methanol stability from 35 to 40% (v/v) after 30 min of pre-incubation. Enhancement in thermostability for the mutant 2DC lipase at 70 °C and 75 °C showed higher half-life at 70 °C and 75 °C for 30 min and 52 min, respectively. The mutant 2DC lipase maintained the same optimum temperature (70 °C) as T1 lipase, while thermally induced unfolding showed the mutant maintained higher rigidity. The kcat/Km values demonstrated a relatively small difference between the T1 lipase (WT) and 2DC lipase (mutant). The kcat/Km (s-1 mM-1) of the T1 and 2DC showed values of 13,043 ± 224 and 13,047 ± 312, respectively. X-ray diffraction of 2DC lipase crystal structure with a resolution of 2.04 Å revealed that the introduced single disulfide bond did not lower initial structural interactions within the residues. Enhanced methanol and thermal stability are suggested to be strongly related to the newly disulfide bridge formation and the enhanced compactness and rigidity of the mutant structure. KEY POINTS: • Protein engineering via rational design revealed relative improved enzymatic performance. • The presence of disulfide bond impacts on the rigidity and structural function of proteins. • X-ray crystallography reveals structural changes accompanying protein modification.

Antimicrobial peptides from Bacillus spp. and strategies to enhance their yield.

Antibiotic resistance is a growing concern that is affecting public health globally. The search for alternative antimicrobial agents has become increasingly important. Antimicrobial peptides (AMPs) produced by Bacillus spp. have emerged as a promising alternative to antibiotics, due to their broad-spectrum antimicrobial activity against resistant pathogens. In this review, we provide an overview of Bacillus-derived AMPs, including their classification into ribosomal (bacteriocins) and non-ribosomal peptides (lipopeptides and polyketides). Additionally, we delve into the molecular mechanisms of AMP production and describe the key biosynthetic gene clusters involved. Despite their potential, the low yield of AMPs produced under normal laboratory conditions remains a challenge to large-scale production. This review thus concludes with a comprehensive summary of recent studies aimed at enhancing the productivity of Bacillus-derived AMPs. In addition to medium optimization and genetic manipulation, various molecular strategies have been explored to increase the production of recombinant antimicrobial peptides (AMPs). These include the selection of appropriate expression systems, the engineering of expression promoters, and metabolic engineering. Bacillus-derived AMPs offer great potential as alternative antimicrobial agents, and this review provides valuable insights on the strategies to enhance their production yield, which may have significant implications for combating antibiotic resistance. KEY POINTS: • Bacillus-derived AMP is a potential alternative therapy for resistant pathogens • Bacillus produces two main classes of AMPs: ribosomal and non-ribosomal peptides • AMP yield can be enhanced using culture optimization and molecular approaches.

Ancestral sequence reconstruction of ancient lipase from family I.3 bacterial lipolytic enzymes.

Family I.3 lipase is distinguished from other families by the amino acid sequence and secretion mechanism. Little is known about the evolutionary process driving these differences. This study attempt to understand how the diverse temperature stabilities of bacterial lipases from family I.3 evolved. To achieve that, eighty-three protein sequences sharing a minimum 30% sequence identity with Antarctic Pseudomonas sp. AMS8 lipase were used to infer phylogenetic tree. Using ancestral sequence reconstruction (ASR) technique, the last universal common ancestor (LUCA) sequence of family I.3 was reconstructed. A gene encoding LUCA was synthesized, cloned and expressed as inclusion bodies in E. coli system. Insoluble form of LUCA was refolded using urea dilution method and then purified using affinity chromatography. The purified LUCA exhibited an optimum temperature and pH at 70 ℃ and 10 respectively. Various metal ions increased or retained the activity of LUCA. LUCA also demonstrated tolerance towards various organic solvents in 25% v/v concentration. The finding from this study could support the understanding on temperature and environment during ancient time. In overall, reconstructed ancestral enzymes have improved physicochemical properties that make them suitable for industrial applications and ASR technique can be employed as a general technique for enzyme engineering.

Bacteria consortia enhanced hydrocarbon degradation of waxy crude oil.

Waxy crude oil is a problem to the oil and gas industry because wax deposition in pipelines reduces the quality of the crude oil. Currently, the industry uses chemicals to solve the problem but it is not environmentally friendly. As an alternative, the biodegradation approach is one of the options. Previously eleven thermophilic bacteria were isolated and exhibited high ability to degrade hydrocarbon up to 70% of waxy crude oil. However, despite the successful study on these single bacteria strains, it is believed that biodegradation of paraffin wax requires more than a single species. Five consortia were developed based on the biodegradation efficiency of 11 bacterial strains. Consortium 3 showed the highest biodegradation (77.77%) with more long-chain alkane degraded throughout the incubation compared to other consortia. Enhancement of hydrocarbon degradation was observed for all consortia especially in long chain alkane (C18-C40). Consortium 3 exhibited higher alkane monooxygenase, alcohol dehydrogenase, lipase, and esterase activities. Moreover, the dominant bacteria in the consortia were determined by denaturing gradient gel electrophoresis (DGGE), which showed the domination of genera Geobacillus, Parageobacillus, and Anoxybacillus. It can be concluded that the bacterial consortia showed higher biodegradation and improved degrading more long-chain hydrocarbon compared to a single isolate.

Membrane fatty acid desaturase: biosynthesis, mechanism, and architecture.

Fatty acid desaturase catalyzes the desaturation reactions by inserting double bonds into the fatty acyl chain, producing unsaturated fatty acids, which play a vital part in the synthesis of polyunsaturated fatty acids. Though soluble fatty acid desaturases have been described extensively in advanced organisms, there are very limited studies of membrane fatty acid desaturases due to their difficulties in producing a sufficient amount of recombinant desaturases. However, the advancement of technology has shown substantial progress towards the development of elucidating crystal structures of membrane fatty acid desaturase, thus, allowing modification of structure to be manipulated. Understanding the structure, mechanism, and biosynthesis of fatty acid desaturase lay a foundation for the potential production of various strategies associated with alteration and modifications of polyunsaturated fatty acids. This manuscript presents the current state of knowledge and understanding about the structure, mechanisms, and biosynthesis of fatty acid desaturase. In addition, the role of unsaturated fatty acid desaturases in health and diseases is also encompassed. This will be useful in understanding the molecular basis and structural protein of fatty acid desaturase that are significant for the advancement of therapeutic strategies associated with the improvement of health status. KEY POINTS: • Current state of knowledge and understanding about the biosynthesis, mechanisms, and structure of fatty acid desaturase. • The role of unsaturated fatty acid desaturase. • The molecular basis and structural protein elucidated the crystal structure of fatty acid desaturase.

Versatility of subtilisin: A review on structure, characteristics, and applications.

Due to its thermostability and high pH compatibility, subtilisin is most known for its role as an additive for detergents in which it is categorized as a serine protease according to MEROPS database. Subtilisin is typically isolated from various bacterial species of the Bacillus genus such as Bacillus subtilis, B. amyloliquefaciens, B. licheniformis, and various other organisms. It is composed of 268-275 amino acid residues and is initially secreted in the precursor form, preprosubtilisin, which is composed of 29-residues signal peptide, 77-residues propeptide, and 275-residues active subtilisin. Subtilisin is known for the presence of high and low affinity calcium binding sites in its structure. Native subtilisin has general properties of thermostability, tolerance to neutral to high pH, broad specificity, and calcium-dependent stability, which contribute to the versatility of subtilisin applicability. Through protein engineering and immobilization technologies, many variants of subtilisin have been generated, which increase the applicability of subtilisin in various industries including detergent, food processing and packaging, synthesis of inhibitory peptides, therapeutic, and waste management applications.

Cold-Active Lipases and Esterases: A Review on Recombinant Overexpression and Other Essential Issues.

Cold environments characterised by diverse temperatures close to or below the water freezing point dominate about 80% of the Earth's biosphere. One of the survival strategies adopted by microorganisms living in cold environments is their expression of cold-active enzymes that enable them to perform an efficient metabolic flux at low temperatures necessary to thrive and reproduce under those constraints. Cold-active enzymes are ideal biocatalysts that can reduce the need for heating procedures and improve industrial processes' quality, sustainability, and cost-effectiveness. Despite their wide applications, their industrial usage is still limited, and the major contributing factor is the lack of complete understanding of their structure and cold adaptation mechanisms. The current review looked at the recombinant overexpression, purification, and recent mechanism of cold adaptation, various approaches for purification, and three-dimensional (3D) crystal structure elucidation of cold-active lipases and esterase.

Thermostable lipases and their dynamics of improved enzymatic properties.

Thermal stability is one of the most desirable characteristics in the search for novel lipases. The search for thermophilic microorganisms for synthesising functional enzyme biocatalysts with the ability to withstand high temperature, and capacity to maintain their native state in extreme conditions opens up new opportunities for their biotechnological applications. Thermophilic organisms are one of the most favoured organisms, whose distinctive characteristics are extremely related to their cellular constituent particularly biologically active proteins. Modifications on the enzyme structure are critical in optimizing the stability of enzyme to thermophilic conditions. Thermostable lipases are one of the most favourable enzymes used in food industries, pharmaceutical field, and actively been studied as potential biocatalyst in biodiesel production and other biotechnology application. Particularly, there is a trade-off between the use of enzymes in high concentration of organic solvents and product generation. Enhancement of the enzyme stability needs to be achieved for them to maintain their enzymatic activity regardless the environment. Various approaches on protein modification applied since decades ago conveyed a better understanding on how to improve the enzymatic properties in thermophilic bacteria. In fact, preliminary approach using advanced computational analysis is practically conducted before any modification is being performed experimentally. Apart from that, isolation of novel extremozymes from various microorganisms are offering great frontier in explaining the crucial native interaction within the molecules which could help in protein engineering. In this review, the thermostability prospect of lipases and the utility of protein engineering insights into achieving functional industrial usefulness at their high temperature habitat are highlighted. Similarly, the underlying thermodynamic and structural basis that defines the forces that stabilize these thermostable lipase is discussed. KEY POINTS: • The dynamics of lipases contributes to their non-covalent interactions and structural stability. • Thermostability can be enhanced by well-established genetic tools for improved kinetic efficiency. • Molecular dynamics greatly provides structure-function insights on thermodynamics of lipase.

Ion-Pair Interaction and Hydrogen Bonds as Main Features of Protein Thermostability in Mutated T1 Recombinant Lipase Originating from Geobacillus zalihae.

A comparative structure analysis between space- and an Earth-grown T1 recombinant lipase from Geobacillus zalihae had shown changes in the formation of hydrogen bonds and ion-pair interactions. Using the space-grown T1 lipase validated structure having incorporated said interactions, the recombinant T1 lipase was re-engineered to determine the changes brought by these interactions to the structure and stability of lipase. To understand the effects of mutation on T1 recombinant lipase, five mutants were developed from the structure of space-grown T1 lipase and biochemically characterized. The results demonstrate an increase in melting temperature up to 77.4 °C and 76.0 °C in E226D and D43E, respectively. Moreover, the mutated lipases D43E and E226D had additional hydrogen bonds and ion-pair interactions in their structures due to the improvement of stability, as observed in a longer half-life and an increased melting temperature. The biophysical study revealed differences in ß-Sheet percentage between less stable (T118N) and other mutants. As a conclusion, the comparative analysis of the tertiary structure and specific residues associated with ion-pair interactions and hydrogen bonds could be significant in revealing the thermostability of an enzyme with industrial importance.

Changes of Thermostability, Organic Solvent, and pH Stability in Geobacillus zalihae HT1 and Its Mutant by Calcium Ion.

Thermostable T1 lipase from Geobacillus zalihae has been crystallized using counter-diffusion method under space and Earth conditions. The comparison of the three-dimensional structures from both crystallized proteins show differences in the formation of hydrogen bond and ion interactions. Hydrogen bond and ion interaction are important in the stabilization of protein structure towards extreme temperature and organic solvents. In this study, the differences of hydrogen bond interactions at position Asp43, Thr118, Glu250, and Asn304 and ion interaction at position Glu226 was chosen to imitate space-grown crystal structure, and the impact of these combined interactions in T1 lipase-mutated structure was studied. Using space-grown T1 lipase structure as a reference, subsequent simultaneous mutation D43E, T118N, E226D, E250L, and N304E was performed on recombinant wild-type T1 lipase (wt-HT1) to generate a quintuple mutant term as 5M mutant lipase. This mutant lipase shared similar characteristics to its wild-type in terms of optimal pH and temperature. The stability of mutant 5M lipase improved significantly in acidic and alkaline pH as compared to wt-HT1. 5M lipase was highly stable in organic solvents such as dimethyl sulfoxide (DMSO), methanol, and n-hexane compared to wt-HT1. Both wild-type and mutant lipases were found highly activated in calcium as compared to other metal ions due to the presence of calcium-binding site for thermostability. The presence of calcium prolonged the half-life of mutant 5M and wt-HT1, and at the same time increased their melting temperature (Tm). The melting temperature of 5M and wt-HT1 lipases increased at 8.4 and 12.1 °C, respectively, in the presence of calcium as compared to those without. Calcium enhanced the stability of mutant 5M in 25% (v/v) DMSO, n-hexane, and n-heptane. The lipase activity of wt-HT1 also increased in 25% (v/v) ethanol, methanol, acetonitrile, n-hexane, and n-heptane in the presence of calcium. The current study showed that the accumulation of amino acid substitutions D43E, T118N, E226D, E250L, and N304E produced highly stable T1 mutant when hydrolyzing oil in selected organic solvents such as DMSO, n-hexane, and n-heptane. It is also believed that calcium ion plays important role in regulating lipase thermostability.

Insight into Improved Thermostability of Cold-Adapted Staphylococcal Lipase by Glycine to Cysteine Mutation.

Thermostability remains one of the most desirable traits in many lipases. Numerous studies have revealed promising strategies to improve thermostability and random mutagenesis often leads to unexpected yet interesting findings in engineering stability. Previously, the thermostability of C-terminal truncated cold-adapted lipase from Staphylococcus epidermidis AT2 (rT-M386) was markedly enhanced by directed evolution. The newly evolved mutant, G210C, demonstrated an optimal temperature shift from 25 to 45 °C and stability up to 50 °C. Interestingly, a cysteine residue was randomly introduced on the loop connecting the two lids and accounted for the only cysteine found in the lipase. We further investigated the structural and mechanistic insights that could possibly cause the significant temperature shift. Both rT-M386 and G210C were modeled and simulated at 25 °C and 50 °C. The results clearly portrayed the effect of cysteine substitution primarily on the lid stability. Comparative molecular dynamics simulation analysis revealed that G210C exhibited greater stability than the wild-type at high temperature simulation. The compactness of the G210C lipase structure increased at 50 °C and resulted in enhanced rigidity hence stability. This observation is supported by the improved and stronger non-covalent interactions formed in the protein structure. Our findings suggest that the introduction of a single cysteine residue at the lid region of cold-adapted lipase may result in unexpected increased in thermostability, thus this approach could serve as one of the thermostabilization strategies in engineering lipase stability.

Polyunsaturated fatty acids in marine bacteria and strategies to enhance their production.

Polyunsaturated fatty acids (PUFAs) play an important role in human diet. Despite the wide-ranging importance and benefits from heart health to brain functions, humans and mammals cannot synthesize PUFAs de novo. The primary sources of PUFA are fish and plants. Due to the increasing concerns associated with food security as well as issues of environmental contaminants in fish oil, there has been considerable interest in the production of polyunsaturated fatty acids from alternative resources which are more sustainable, safer, and economical. For instance, marine bacteria, particularly the genus of Shewanella, Photobacterium, Colwellia, Moritella, Psychromonas, Vibrio, and Alteromonas, are found to be one among the major microbial producers of polyunsaturated fatty acids. Recent developments in the area with a focus on the production of polyunsaturated fatty acids from marine bacteria as well as the metabolic engineering strategies for the improvement of PUFA production are discussed.

The biology and the importance of Photobacterium species.

Photobacterium species are Gram-negative coccobacilli which are distributed in marine habitats worldwide. Some species are unique because of their capability to produce luminescence. Taxonomically, about 23 species and 2 subspecies are validated to date. Genomes from a few Photobacterium spp. have been sequenced and studied. They are considered a special group of bacteria because some species are capable of producing essential polyunsaturated fatty acids, antibacterial compounds, lipases, esterases and asparaginases. They are also used as biosensors in food and environmental monitoring and detectors of drown victim, as well as an important symbiont.

Improving the Efficiency of New Automatic Dishwashing Detergent Formulation by Addition of Thermostable Lipase, Protease and Amylase.

The use of T1 lipase in automatic dishwashing detergent (ADD) is well established, but efficiency in hard water is very low. A new enzymatic environmentally-friendly dishwashing was formulated to be efficient in both soft and hard water. Thermostable enzymes such as T1 lipase from Geobacillus strain T1, Rand protease from Bacillussubtilis strain Rand, and Maltogenic amylase from Geobacillus sp. SK70 were produced and evaluated for an automatic dishwashing detergent formulation. The components of the new ADD were optimized for compatibility with these three enzymes. In compatibility tests of the enzymes with different components, several criteria were considered. The enzymes were mostly stable in non-ionic surfactants, especially polyhydric alcohols, Glucopon UP 600, and in a mixture of sodium carbonate and glycine (30:70) buffer at a pH of 9.25. Sodium polyacrylate and sodium citrate were used in the ADD formulation as a dispersing agent and a builder, respectively. Dishwashing performance of the formulated ADDs was evaluated in terms of percent of soil removed using the Leenert's Improved Detergency Tester. The results showed that the combination of different hydrolysis enzymes could improve the washing efficiency of formulated ADD compared to the commercial ADD "Finish" at 40 and 50 C.

Molecular Dynamic Simulation of Space and Earth-Grown Crystal Structures of Thermostable T1 Lipase Geobacillus zalihae Revealed a Better Structure.

Less sedimentation and convection in a microgravity environment has become a well-suited condition for growing high quality protein crystals. Thermostable T1 lipase derived from bacterium Geobacilluszalihae has been crystallized using the counter diffusion method under space and earth conditions. Preliminary study using YASARA molecular modeling structure program for both structures showed differences in number of hydrogen bond, ionic interaction, and conformation. The space-grown crystal structure contains more hydrogen bonds as compared with the earth-grown crystal structure. A molecular dynamics simulation study was used to provide insight on the fluctuations and conformational changes of both T1 lipase structures. The analysis of root mean square deviation (RMSD), radius of gyration, and root mean square fluctuation (RMSF) showed that space-grown structure is more stable than the earth-grown structure. Space-structure also showed more hydrogen bonds and ion interactions compared to the earth-grown structure. Further analysis also revealed that the space-grown structure has long-lived interactions, hence it is considered as the more stable structure. This study provides the conformational dynamics of T1 lipase crystal structure grown in space and earth condition.

Cloning, expression and characterization of a novel cold­adapted GDSL family esterase from Photobacterium sp. strain J15.

The gene encoding for a novel cold-adapted enzyme from family II of bacterial classification (GDSL family) was cloned from the genomic DNA of Photobacterium sp. strain J15 in an Escherichia coli system, yielding a recombinant 36 kDa J15 GDSL esterase which was purified in two steps with a final yield and purification of 38.6 and 15.3 respectively. Characterization of the biochemical properties showed the J15 GDSL esterase had maximum activity at 20 °C and pH 8.0, was stable at 10 °C for 3 h and retained 50 % of its activity after a 6 h incubation at 10 °C. The enzyme was activated by Tween-20, -60 and Triton-X100 and inhibited by 1 mM Sodium dodecyl sulphate (SDS), while ß-mercaptoethanol and Dithiothreitol (DTT) enhanced activity by 4.3 and 5.4 fold respectively. These results showed the J15 GDSL esterase was a novel cold-adapted enzyme from family II of lipolytic enzymes. A structural model constructed using autotransporter EstA from Pseudomonas aeruginosa as a template revealed the presence of a typical catalytic triad consisting of a serine, aspartate, and histidine which was verified with site directed mutagenesis on active serine.

Adaptational properties and applications of cold-active lipases from psychrophilic bacteria.

Psychrophilic microorganisms are cold-adapted with distinct properties from other thermal classes thriving in cold conditions in large areas of the earth's cold environment. Maintenance of functional membranes, evolving cold-adapted enzymes and synthesizing a range of structural features are basic adaptive strategies of psychrophiles. Among the cold-evolved enzymes are the cold-active lipases, a group of microbial lipases with inherent stability-activity-flexibility property that have engaged the interest of researchers over the years. Current knowledge regarding these cold-evolved enzymes in psychrophilic bacteria proves a display of high catalytic efficiency with low thermal stability, which is a differentiating feature with that of their mesophilic and thermophilic counterparts. Improvement strategies of their adaptive structural features have significantly benefited the enzyme industry. Based on their homogeneity and purity, molecular characterizations of these enzymes have been successful and their properties make them unique biocatalysts for various industrial and biotechnological applications. Although, strong association of lipopolysaccharides from Antarctic microorganisms with lipid hydrolases pose a challenge in their purification, heterologous expression of the cold-adapted lipases with affinity tags simplifies purification with higher yield. The review discusses these cold-evolved lipases from bacteria and their peculiar properties, in addition to their potential biotechnological and industrial applications.