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Lipases are glycerol ester hydrolases (E.C. 3.1.1.3) that catalyze the hydrolysis of triacylglycerols to free fatty acids and glycerol. They resemble esterases (E.C. 3.1.1.1) in catalytic activity but differ in that substrates. True lipases prefer water-insoluble fats containing medium-to long-chain fatty acids. Lipases are used extensively in the detergent, food, dairy, pulp, and pharmaceutical industries due to their high productivity and diversity, such as substrate specificity, stability in organic solvents, and high degree of regioselectivity [1].Bacterial lipases are classified into eight families based on amino acid sequence homology [2]. Family I lipases, called true lipases, are large group which is further divided into 6 subfamilies. They possess the pentapeptide Gly-Xaa-Ser-Xaa-Gly (GxSxG) motif with the active site serine situated near the center of the conserved sequence [2,3]. Most of the bacterial lipases from Bacillus and Staphyloccocus species belongs to this family. The enzymes grouped in family II do not exhibit the conventional GxSxG motif but rather display a Gly-Asp-Ser-Leu (GDSL) motif containing the active site serine residue. The GDSL motif localized in near N-terminus of amino acid sequence which is compared to GxSxG motif conserved in center of the sequence [4]. GDSL lipases represent the lipolytic activities with multifunctional properties and broad substrate specificity [5,6]. Furthermore, a subgroup of this GDSL family was classified as the SGNH hydrolase superfamily, with four conserved residues Ser, Gly, Asn and His in four conserved blocks I, II, III, and V [6]. While the SGNH hydrolases are well known in eukaryotic organisms, the isolation and characterization of SGNH hydrolases from bacteria remain to be limited [7]. In bacteria, GDSL motif enzymes are generally known as esterase type which has preference to short chain fatty acids [8][9][10][11]. To date, there have been few reports of GDSL family lipases in bacteria [12,13]. One example is a GDSL lipase from Mycobacterium tuberculosis and it was known to be actively involved in the intracellular survival during the nutritive stress conditions [12].Geobacillus species, which belongs to thermophilic Gram-positive spore-forming bacteria that can grow over a range of 45-75 o C, are of interest for biotechnology field as source of thermostable enzymes. Also, Geobacillus species are known to have potential availability for digesters of lignocellulose, hydrocarbons bioremediators, biofuel producers, cellular factories for heterologous expression of enzymes because of their structural and functional stability in extreme environments [14][15][16][17]. Several lipases which belong to family I have been reported from this species [18,19]. A number of family I and II esterases from this species have been characterized [20,21] Two putative genes, lip29 and est29, encoding lipolytic enzymes from the thermophilic bacterium Geobacillus thermocatenulatus KCTC 3921 were cloned and overexpressed in Escherichia coli. The recombinant Lip29 and Est...

The Crenarchaeon Sulfolobus acidocaldarius has been described to synthesize trehalose via the maltooligosyltrehalose synthase (TreY) and maltooligosyltrehalose trehalohydrolase (TreZ) pathway and the trehalose glycosyltransferring synthase (TreT) pathway has been predicted. Deletion mutant analysis of single and double deletion strains of ΔtreY and ΔtreT in S. acidocaldarius revealed that next to these two pathways a third, novel trehalose biosynthesis pathway is operative in vivo: the trehalose-6-phosphate (T6P) synthase/T6P phosphatase (TPS/TPP) pathway. In contrast to known TPS proteins, which belong to the GT20 family, the S. acidocaldarius TPS belongs to the GT4 family establishing a new function within this group of enzymes. This novel GT4-like TPS turned out to be mainly present in the Sulfolobales. The ΔtreY/ΔtreT/Δtps triple mutant of S. acidocaldarius lacking the ability to synthesize trehalose showed no altered phenotype under standard conditions or heat stress, but was unable to grow under salt stress. Accordingly, in the wild type strain a significant increase of intracellular trehalose formation was observed under salt stress. Quantitative real-time PCR showed a salt stress mediated induction of all three trehalose synthesizing pathways. This demonstrates in Archaea that trehalose plays an essential role for growth under high salt conditions. Importance The metabolism and function of trehalose as compatible solute was not well understood in Archaea. This combined genetic and enzymatic approach at the interface of microbiology, physiology and microbial ecology gives important insights into survival under stress, adaptation to extreme environments, and the role of compatible solutes in Archaea. Here we unravelled the complexity of trehalose metabolism and present a comprehensive study on trehalose function in stress response in S. acidocaldarius. This sheds light on the general microbiology and the fascinating metabolic repertoire of Archaea involving many novel biocatalysts such as glycosyltransferases with great potential in biotechnology.

Saeu-jeotgal, a Korean fermented shrimp food, is commonly used as an ingredient for making kimchi and other side dishes. The high salinity of the jeotgal contributes to its flavor and inhibits the growth of food spoilage microorganisms. Interestingly, Staphylococcus saprophyticus was discovered to be capable of growth even after treatment with 20% NaCl. To elucidate the tolerance mechanism, a genome-wide gene expression of S. saprophyticus against 0%, 10%, and 20% NaCl was investigated by RNA sequencing. A total of 831, 1314, and 1028 differentially expressed genes (DEGs) were identified in the 0% vs. 10%, 0% vs. 20%, and 10% vs. 20% NaCl comparisons, respectively. The Clusters of Orthologous Groups analysis revealed that the DEGs were involved in amino acid transport and metabolism, transcription, and inorganic ion transport and metabolism. The functional enrichment analysis showed that the expression of the genes encoding mechanosensitive ion channels, sodium/proton antiporters, and betaine/carnitine/choline transporter family proteins was downregulated, whereas the expression of the genes encoding universal stress proteins and enzymes for glutamate, glycine, and alanine synthesis was upregulated. Therefore, these findings suggest that the S. saprophyticus isolated from the saeu-jeotgal utilizes different molecular strategies for halotolerance, with glutamate as the key molecule.

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