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The stability and activity of lipase in organic media are important parameters in determining how quickly biocatalysis proceeds. This study aimed to examine the effects of two commonly used alcohols in industrial applications, methanol (MtOH) and ethanol (EtOH) on the conformational stability and catalytic activity of G210C lipase, a laboratory-evolved mutant of Staphylococcus epidermidis AT2 lipase. Simulation studies were performed using an open-form predicted structure under 30, 40 and 50 of MtOH and EtOH at 25‰°C and 45‰°C. The overall enzyme structure becomes more flexible with increasing concentration of MtOH and exhibited the highest flexibility in 40 EtOH. In EtOH, the movement of the lid was found to be temperature-dependent with a noticeable shift in the lid position at 45‰°C. Lid opening was evidenced at 50 of MtOH and EtOH which was supported by the increase in SASA of hydrophobic residues of the lid and catalytic triad. The active site remained mostly intact. An open-closed lid transition was observed when the structure was re-simulated in water. Experimental evaluation of the lipase stability showed that the half-life reduced when the enzyme was treated with 40 (v/v) and 50 (v/v) of EtOH and MtOH respectively. The finding implies that a high concentration of alcohol and elevated temperature can induce the lid opening of lipase which could be essential for the activation of the enzyme, provided that the catalytic performance in the active site is not compromised.

Invasive candidiasis caused by the pathogenic Candida yeast species has resulted in elevating global mortality. The pathogenicity of Candida spp. is not only originated from its primary invasive yeast-to-hyphal transition; virulence factors (transcription factors, adhesins, invasins, and enzymes), biofilm, antifungal drug resistance, stress tolerance, and metabolic adaptation have also contributed to a greater clinical burden. However, the current research theme in fungal pathogenicity could hardly be delineated with the increasing research output. Therefore, our study analyzed the research trends in Candida pathogenesis over the past 37 years via a bibliometric approach against the Scopus and Web of Science databases. Based on the 3993 unique documents retrieved, significant international collaborations among researchers were observed, especially between Germany (Bernhard Hube) and the UK (Julian Naglik) whose focuses are on Candida proteinases, adhesins, and Candidalysin. The prominent researchers (Neils Gow, Alistair Brown and Frank Odds) at the University of Exeter and University of Aberdeen (second top performing affiliation), UK contribute significantly to the mechanisms of Candida adaptation, tolerance, and stress response. However, the science mapping of co-citation analysis performed herein could not identify a hub representative of subsequent work since the clusters were semi-redundant. The co-word analysis that was otherwise adopted, revealed three research clusters; the cluster-based thematic analyses indicated the severeness of Candida biofilm and antifungal resistance as well as the elevating trend on molecular mechanism elucidation for drug screening and repurposing. Importantly, the in vivo pathogen adaptation and interactions with hosts are crucial for potential vaccine development.

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.

Over the last decade, an escalating number of reported cases of leptospirosis in Malaysia has resulted in a significant number of deaths. Leptospirosis is caused by Leptospira sp., which is disseminated by soil and water. Due to the absolute dependence of microbes on this biosynthesis pathway coupled with its non-existence in humans, the structural analysis of riboflavin synthase as a potential target for the development of antimicrobial agents is very significant. Riboflavin synthase (E.C 2.5.1.9) is an important enzyme that catalyzes the last stage of riboflavin biosynthesis. Riboflavin synthase from the locally isolated pathogenic bacterium Leptospira kmetyi was cloned and expressed in Escherichia coli. The protein was purified and crystallized for X-ray analysis. The crystal structure of riboflavin synthase from L. kmetyi has been determined at 3.2 Å and belonged to the orthorhombic space group P212121, with an R-factor of 21.8%. The calculated Matthews coefficient (VM) was 2.94 Å3Da−1 with a solvent content of 58.24 %. There are three molecules present in the asymmetric unit. The structure was analyzed with potential inhibitors via molecular docking and molecular dynamics simulation. The complex with compound 1 had the best possible interaction and stability for a 100 ns simulation. The existence of this crystal structure enables its structural properties to be unveiled and facilitates the application of structure-based drug discovery approach. This study sheds light on the development of new potential antibacterial drugs that may be beneficial for the treatment of leptospirosis or other infectious diseases associated with it.

Lightning is a naturally occurring phenomenon that involves a sudden electrostatic discharge caused by an imbalance between electrically charged cloud regions. Although lightning is visibly amazing, its impact can be dangerous and damaging, which many studies have carried out lightning-generated electric field measurements to assess the electrical discharge features. This study conducted the lightning-generated electric field measurement on the College of Engineering building rooftop at UNITEN from August 2019 to March 2020. A total of 115 negative lightning return strokes waveforms were recorded using a parallel plate antenna. A comparison was made between the data measured in the tropical and non-tropical regions, such as UTM, UPM, Sweden, USA, and Germany, in terms of the characteristic, mainly on the negative return strokes parameters. It was observed that data measured in the same region, either tropical or non-tropical, were consistent or almost similar. On the contrary, the results indicated a significant difference between these two regions on the negative return strokes parameters characteristics. The zero-to-peak and fast transition 10–90% rise time, as well as width dE/dt pulse at half peak value in the tropical region, were observed higher than the non-tropical region. Meanwhile, the zero-crossing time and slow front amplitude relative to the peak in the non-tropical region were averagely longer as compared to the tropical region. Therefore, dissimilarities in the characteristics of negative return stroke parameters could be attributed to the variation in the meteorological conditions, geographical locations, and climatic affection.

In modern urban society, plastic has become a major component in many applications. Despite their usefulness, plastics are known for their alarming resistance to biodegradation and therefore pose a huge burden on the environment. The surge in plastic waste has led to the development of biodegradable plastics. Biodegradable plastics are perceived to be one of the sustainable ways to reduce plastic wastes. Among the biodegradable plastics, polylactic acid (PLA), a bio-based plastic, has been widely used for diverse applications. PLA however, is known to be less susceptible to biodegradation and requires longer time to degrade in the natural environment compared to other aliphatic polyesters. Therefore, identification of PLA-degraders is the current focus of research. Potential PLA-degraders have been isolated from various sources, but the number is still limited. Considering temperature as one of the important factors in biodegradation, in this review, we categorize PLA degraders into three groups; thermophilic, mesophilic and psychrophilic, and discuss their relevance to PLA biodegradation. Enzymes associated with PLA degradation are described according to their temperature preference. One of the current limitations is the structural insights into the catalytic mechanism. Discovery of more PLA-degraders and understanding of the mechanism could aid in designing strategies to accelerate the biodegradation process.

Candidiasis is a fungal infection caused by Candida spp. especially Candida albicans, C. glabrata, C. parapsilosis and C. tropicalis. Although the medicinal therapeutic strategies have rapidly improved, the mortality rate as candidiasis has continuously increased. The secreted and membrane-bound virulence factors (VFs) are responsible for fungal invasion, damage and translocation through the host enterocytes besides the evasion from host immune system. VFs such as agglutinin-like sequences (Als), heat shock protein 70, phospholipases, secreted aspartyl proteinases (Sap), lipases, enolases and phytases are mostly hydrolases which degrade or interact with the enterocyte membrane components. Candidalysin, however, acts as a peptide toxin to induce necrotic cell lysis. To date, structural studies of the VFs remain underexplored, hindering their functional analyses. Among the VFs, only Sap and Als have their structures deposited in Protein Data Bank (PDB). Therefore, this review scrutinizes the mechanisms of these VFs by discussing the VF-deficient studies of several Candida spp. and their abilities to produce these VFs. Nonetheless, their latest reported sequential and structural analyses are discussed to impart a wider perception of the host-pathogen interactions and potential vaccine or antifungal drug targets. This review signifies that more VFs structural investigations and mining in the emerging Candida spp. are required to decipher their pathogenicity and virulence mechanisms compared to the prominent C. albicans.

Aminopeptidase P (APPro, E.C 3.4.11.9) cleaves N-terminal amino acids from peptides and proteins where the penultimate residue is proline. This metal-ion-dependent enzyme shares a similar fold, catalytic mechanism, and substrate specificity with methionine aminopeptidase and prolidase. It adopts a canonical pita bread fold that serves as a structural basis for the metal-dependent catalysis and assembles as a tetramer in crystals. Similar to other metalloaminopeptidase, APPro requires metal ions for its maximal enzymatic activity, with manganese being the most preferred cation. Microbial aminopeptidase possesses unique characteristics compared with aminopeptidase from other sources, making it a great industrial enzyme for various applications. This review provides a summary of recent progress in the study of the structure and function of aminopeptidase P and describes its various applications in different industries as well as its significance in the environment.

Microbial lipases represent one of the most important groups of biotechnological biocatalysts. However, the high-level production of lipases requires an understanding of the molecular mechanisms of gene expression, folding, and secretion processes. Stable, selective, and productive lipase is essential for modern chemical industries, as most lipases cannot work in different process conditions. However, the screening and isolation of a new lipase with desired and specific properties would be time consuming, and costly, so researchers typically modify an available lipase with a certain potential for minimizing cost. Improving enzyme properties is associated with altering the enzymatic structure by changing one or several amino acids in the protein sequence. This review detailed the main sources, classification, structural properties, and mutagenic approaches, such as rational design (site direct mutagenesis, iterative saturation mutagenesis) and direct evolution (error prone PCR, DNA shuffling), for achieving modification goals. Here, both techniques were reviewed, with different results for lipase engineering, with a particular focus on improving or changing lipase specificity. Changing the amino acid sequences of the binding pocket or lid region of the lipase led to remarkable enzyme substrate specificity and enantioselectivity improvement. Site-directed mutagenesis is one of the appropriate methods to alter the enzyme sequence, as compared to random mutagenesis, such as error-prone PCR. This contribution has summarized and evaluated several experimental studies on modifying the substrate specificity of lipases.

Citrobacter sp. S-77 [NiFe]-hydrogenase harbors a standard [4Fe–4S] cluster proximal to the Ni–Fe active site. The presence of relocatable water molecules and a flexible aspartate enables the [4Fe–4S] to display redox-dependent conformational changes. These structural features are proposed to be the key aspects that protect the active site from O2 attack.

Structural comparison of wild-type F1 protease and thermophilic homologues leads to hypothesis about the role of ion pairs for the thermostability of the enzymes. Higher thermostability was found to correlate with an increased number of the residues involved in ion pairs or ion pair’s networks. Therefore, W200R protease is a mutant designed to have three new additional ion pairs compared to the F1 protease. This designed mutant underwent substitution of Trp (Tryptophan) at position 200 with Arg (Arginine). Both of protease and W200R were cloned into pGEX-4T1 vector and transformed into E. coli BL21 (DE3)pLysS. For the purpose of characterization, F1 protease was purified by using Glutathione Sepharose resin and heat treatment whereby W200R was purified in three steps; Glutathione Sepharose resin, ion exchange by using Q-Sepharose Fast Flow resin and Phenyl Sepharose High Performance. The last product of purification for F1 protease and W200R was confirmed to be the mature form which is approximately 27 kDa. Characterization of W200R’s thermostability was done in comparison with F1 protease. The optimum temperature of W200R protease in the presence and absence of 2 mM CaCl2 was 75°C. Meanwhile, F1 protease exhibited a lower temperature optimum which is 70°C with the presence and absence of Ca2+. In term of thermostability in the presence of 2 mM Ca2+, the activity of W200R and F1 protease retained 100% of its activity between 50°C to 80°C and 50°C to 65°C, respectively. Ca2+ affected both enzymes by increasing and stabilizing the protease activity. The optimum pH of W200R was at pH 8 and the activity was stable in pH ranging from pH 7.0-10. The activity of W200R retained more than 100% of its activity compared to the control in the presence of miscible organic solvents log P < 2 such as pyridine, ethanol, methanol and DMSO. In comparison with the wild type, stability in the presence of immiscible organic solvents log P > 2 such as n- heptanes, hexane and benzene was