Levansucrase/Invertase. This Pfam family consists of the glycosyl hydrolase 68 family, including several bacterial levansucrase enzymes, and invertase from zymomonas.

Glycosyl hydrolase family 68, includes levansucrase, beta-fructofuranosidase and inulosucrase. Glycosyl hydrolase family 68 (GH68) consists of frucosyltransferases (FTFs) that include levansucrase (EC 2.4.1.10), beta-fructofuranosidase (EC 3.2.1.26) and inulosucrase (EC 2.4.1.9), all of which use sucrose as their preferential donor substrate. Levansucrase, also known as beta-D-fructofuranosyl transferase, catalyzes the transfer of the sucrose fructosyl moiety to a growing levan chain. Similarly, inulosucrase catalyzes long inulin-type of fructans, and beta-fructofuranosidases create fructooligosaccharides (FOS). However, in the absence of high fructan/sucrose ratio, some GH68 enzymes can also use fructan as donor substrate. GH68 retaining enzymes (i.e. they retain the configuration at anomeric carbon atom of the substrate) catalyze hydrolysis in two steps involving a covalent glycosyl enzyme intermediate: an aspartate located close to the N-terminus acts as the catalytic nucleophile and a glutamate acts as the general acid/base; a conserved aspartate residue in the Arg-Asp-Pro (RDP) motif stabilizes the transition state. A common structural feature of all these enzymes is a 5-bladed beta-propeller domain, similar to GH43, that contains the catalytic acid and catalytic base. A long V-shaped groove, partially enclosed at one end, forms a single extended substrate-binding surface across the face of the propeller. Biotechnological applications of these enzymes include use of inulin in inexpensive production of rich fructose syrups as well as use of FOS as health-promoting pre-biotics.

Glycosyl hydrolase families 32 and 68, which form the clan GH-J. This glycosyl hydrolase family clan J (according to carbohydrate-active enzymes database (CAZY)) includes family 32 (GH32) and 68 (GH68). GH32 enzymes include invertase (EC 3.2.1.26) and other other fructofuranosidases such as inulinase (EC 3.2.1.7), exo-inulinase (EC 3.2.1.80), levanase (EC 3.2.1.65), and transfructosidases such sucrose:sucrose 1-fructosyltransferase (EC 2.4.1.99), fructan:fructan 1-fructosyltransferase (EC 2.4.1.100), sucrose:fructan 6-fructosyltransferase (EC 2.4.1.10), fructan:fructan 6G-fructosyltransferase (EC 2.4.1.243) and levan fructosyltransferases (EC 2.4.1.-). The GH68 family consists of frucosyltransferases (FTFs) that include levansucrase (EC 2.4.1.10, also known as beta-D-fructofuranosyl transferase), beta-fructofuranosidase (EC 3.2.1.26) and inulosucrase (EC 2.4.1.9). GH32 and GH68 family enzymes are retaining enzymes (i.e. they retain the configuration at anomeric carbon atom of the substrate) and catalyze hydrolysis in two steps involving a covalent glycosyl enzyme intermediate: an aspartate located close to the N-terminus acts as the catalytic nucleophile and a glutamate acts as the general acid/base; a conserved aspartate residue in the Arg-Asp-Pro (RDP) motif stabilizes the transition state. A common structural feature of all these enzymes is a 5-bladed beta-propeller domain, similar to GH43, that contains the catalytic acid and catalytic base. A long V-shaped groove, partially enclosed at one end, forms a single extended substrate-binding surface across the face of the propeller.

Glycosyl hydrolase family 43 such as arabinan endo-1 5-alpha-L-arabinosidase. This glycosyl hydrolase family 43 (GH43) subgroup includes mostly enzymes with endo-alpha-L-arabinanase (ABN; EC 3.2.1.99) activity. These are inverting enzymes (i.e. they invert the stereochemistry of the anomeric carbon atom of the substrate) that have an aspartate as the catalytic general base, a glutamate as the catalytic general acid and another aspartate that is responsible for pKa modulation and orienting the catalytic acid. The GH43 ABN enzymes hydrolyze alpha-1,5-L-arabinofuranoside linkages. These arabinan-degrading enzymes are important in the food industry for efficient production of L-arabinose from agricultural waste; L-arabinose is often used as a bioactive sweetener. A common structural feature of GH43 enzymes is a 5-bladed beta-propeller domain that contains the catalytic acid and catalytic base. A long V-shaped groove, partially enclosed at one end, forms a single extended substrate-binding surface across the face of the propeller.

Glycosyl hydrolase family 32, such as the putative glycoside hydrolase Escherichia coli Aec43 (FosGH2). This glycosyl hydrolase family 32 (GH32) subgroup includes Escherichia coli strain BEN2908 putative glycoside hydrolase Aec43 (FosGH2). GH32 enzymes cleave sucrose into fructose and glucose via beta-fructofuranosidase activity, producing invert sugar that is a mixture of dextrorotatory D-glucose and levorotatory D-fructose, thus named invertase (EC 3.2.1.26). GH32 family also contains other fructofuranosidases such as inulinase (EC 3.2.1.7), exo-inulinase (EC 3.2.1.80), levanase (EC 3.2.1.65), and transfructosidases such sucrose:sucrose 1-fructosyltransferase (EC 2.4.1.99), fructan:fructan 1-fructosyltransferase (EC 2.4.1.100), sucrose:fructan 6-fructosyltransferase (EC 2.4.1.10), fructan:fructan 6G-fructosyltransferase (EC 2.4.1.243) and levan fructosyltransferases (EC 2.4.1.-). These retaining enzymes (i.e. they retain the configuration at anomeric carbon atom of the substrate) catalyze hydrolysis in two steps involving a covalent glycosyl enzyme intermediate: an aspartate located close to the N-terminus acts as the catalytic nucleophile and a glutamate acts as the general acid/base; a conserved aspartate residue in the Arg-Asp-Pro (RDP) motif stabilizes the transition state. These enzymes are predicted to display a 5-fold beta-propeller fold as found for GH43 and CH68. The breakdown of sucrose is widely used as a carbon or energy source by bacteria, fungi, and plants. Invertase is used commercially in the confectionery industry, since fructose has a sweeter taste than sucrose and a lower tendency to crystallize.

ChainA, Levansucrase [Bacillus subtilis],3BYN_A Chain A, Levansucrase [Bacillus subtilis]

ChainA, Levansucrase [Bacillus subtilis]

ChainA, Levansucrase [Bacillus subtilis]

Crystalstructure of Levansucrase from Bacillus subtilis mutant S164A at 2.6 A [Bacillus subtilis],6PWQ_B Crystal structure of Levansucrase from Bacillus subtilis mutant S164A at 2.6 A [Bacillus subtilis]

Crystalstructure of Bacillus subtilis levansucrase [Bacillus subtilis]

Inactive levansucrase OS=Geobacillus stearothermophilus OX=1422 GN=sacB PE=1 SV=1

Levansucrase OS=Bacillus subtilis (strain 168) OX=224308 GN=sacB PE=1 SV=1

Levansucrase OS=Bacillus amyloliquefaciens OX=1390 GN=sacB PE=2 SV=1

Levansucrase OS=Fructilactobacillus sanfranciscensis OX=1625 GN=levS PE=1 SV=1

Levansucrase OS=Streptococcus mutans serotype c (strain ATCC 700610 / UA159) OX=210007 GN=ftf PE=3 SV=2