United States of America, As Represented by the Secretary of Agriculture

The United States of America As Represented by the Secretary of

Board of Trustees of Western Illinois Univ.

Gregory L Cote

Timothy D Leathers

Christopher D Skory

Herbert A Wyckoff

Melinda S Nunnally

G Thomas Hayman

Peter Biely

Julious L Willett

Victoria L Finkenstadt

Ryan Cormier

Scott M Holt

Provided herein are compositions and methods for the synthesis of the trisaccharide, isomelezitose, using genetically modified glucansucrase enzymes from representative microorganisms, including lactic acid bacteria such as. Various modified enzymes are detailed, increasing isomelezitose yields and provide the foundation for large-scale production of isomelezitose for food, industrial and biomedical applications.

Modified enzymes for producing increased isomelezitose

Nanoparticles composed of water insoluble glucans and films composed of tile above nanoparticles, in addition, there is disclosed a process for producing nanoparticles composed of water insoluble glucans. Furthermore, surfactant stabilized nanoparticles composed of water insoluble glucans and films composed of the above nanoparticles. In addition, a process for producing surfactant stabilized nanoparticles composed of water insoluble glucans.

Nanoparticles and films composed of water-insoluble glucan

Disclosed is a genetically modified enzyme belonging to glycosyltransferases type of glucansucrase comprising at least one mutation at positionof said enzyme, wherein modified enzyme is capable to producing a glucan polymer.

Modified glucansucrase and related methods

Disclosed is a method for inhibiting corrosion on corrosion-sensitive metal with a bacterial exopolysaccharide. Specifically, exopolysaccharides precipitated from NRRL bacterial strains B-1254, B-1355, B-1498, and B-1500 coated on low carbon steel alloy confers anti-corrosion coating to corrosion sensitive metals. Preferably with the coating, the corrosion rate for coated metal is less than 0.4 milli-inch per year.

Anti-corrosion coating utilizing bacterial precipitated exopolysaccharides

Four newspp. isolates (NRRL 21966, NRRL 21967, NRRL 21968, and NRRL 21969) are capable of essentially quantitative conversion of native alternan to a polymeric modified form having a lower apparent molecular weight than native alternan. A fifth isolate (NRRL 30489) obtained from a survey of deposited organisms and classified as ahas the same ability. The modified alternan has rheological properties similar to ultrasonicated alternan and is produced without the expense of ultrasonication. It would have utility as a substitute for gum arabic for uses such as bulking agents and extenders in foods and cosmetics.

Modified alternan

Oligosaccharides produced by an alternansucrase enzyme catalyzed reaction of sucrose with an acceptor oligosaccharide are effective as prebiotics for controlling enteric bacterial pathogens. Populations of enteropathogenic bacteria may be substantially reduced or inhibited by treatment of an animal with a composition comprising one or more of these oligosaccharides in an amount effective to promote the growth of beneficial bacteria. The method is particularly effective for the control ofspecies, enteropathogenic, and

Prebiotic oligosaccharides via alternansucrase acceptor reactions

Four newspp. isolates (NRRL 21966, NRRL 21967, NRRL 21968, and NRRL 21969) are capable of essentially quantitative conversion of native alternan to a polymeric modified form having a lower apparent molecular weight than native alternan. A fifth isolate (NRRL 30489) obtained from a survey of deposited organisms and classified as asubgenushas the same ability. The modified alternan has rheological properties similar to ultrasonicated alternan and is produced without the expense of ultrasonication. It would have utility as a substitute for gum arabic for uses such as bulking agents and extenders in foods and cosmetics.

The cyclic tetrasaccharide, cyclo&lcub;-6)-&agr;-D-Glcp-(1,3)-&agr;-D-Glcp-(1,6)-&agr;-D-Glcp-(1,3)-&agr;-D-Glcp-(1-&rcub;, may be produced by alternanase hydrolysis of complex carbohydrates other than alternan. Panose, pullulan, &agr;-D-Glcp-(1,6)-&agr;-D-Glcp-(1,3)-D-Glc, and D-glucans having alternating &agr;-(1,6) and &agr;-(1,4) linkages, are all hydrolyzed by alternanase to produce this cyclic tetrasaccharide. In this process, the cyclic tetrasaccharide is produced by contacting a solution of one or more of the above-mentioned complex carbohydrates with an amount of alternanase under conditions effective for activity of the enzyme. The substrate panose used in the reaction may be produced from a variety of polysaccharides or oligosaccharides, including starch, maltose, maltodextrins, pullulan, and mixtures thereof.

Production of cyclic alternan tetrasaccharides from oligosaccharide substrates

Four new Penicillium spp. isolates (NRRL 21966, NRRL 21967, NRRL 21968, and NRRL 21969) are capable of essentially quantitative conversion of native alternan to a polymeric modified form having a lower apparent molecular weight than native alternan. A fifth isolate (NRRL 30489) obtained from a survey of deposited organisms and classified as a Penicillium subgenus Biverticillium has the same ability. The modified alternan has rheological properties similar to ultrasonicated alternan and is produced without the expense of ultrasonication. It would have utility as a substitute for gum arabic for uses such as bulking agents and extenders in foods and cosmetics.

Penicillium isolates for modifying alternan

A new enzyme, alternanase, which is effective for the endo-hydrolytic cleavage of alternan, producing a thinned composition of low molecular weight fractions which exhibit reduced viscosity and increased solubility relative to native alternan, is described. The enzyme is produced and secreted extracellularly by a plurality of novel bacteria isolated from soil. One of the fractions present in the thinned alternan resulting from hydrolysis with alternanase is a the cyclic tetrasaccharide, cyclo{-6)-.alpha.-D-Glcp-(1,3)-.alpha.-D-Glcp-(1,6)-.alpha.-D-Glcp-(1,3)-. alpha.-D-Glcp-(1-}. A novel method for isolating strains of microorganisms which produce endo-.alpha.-D-glucanases such as alternanase effective for the endo-hydrolytic cleavage or thinning of alternan is also described. Cultures of the subject strains are contacted with a test substrate of alternan coupled to a detectable indicator. Detection of released indicator provides an indication of endo-.alpha.-D-glucanase activity.

Bacteria and enzymes for production of alternant fragments

A rapid screening procedure to select microorganism strains that produce a high proportion of alternan to dextan and a high proportion of alternansucrase to dextransucrase is described. Strains selected by the method are useful to produce a high proportion of alternansucrase to dextransucrase and to produce a high proportion of alternan to dextran, either fermentatively by cultivation of the strains on sucrose, or enzymatically by using alternansucrase produced by the strains of the invention. The selected strains can also be used to obtain microorganism strains having further improvements.

Rapid screening method to select microorganism strains that produce a

Bacteria and enzymes for production of alternan fragments

A rapid screening procedure to select microorganism strains that produce a high proportion of alternan to dextran and a high proportion of alternansucrase to dextransucrase is described. Strains selected by the method are useful to produce a high proportion of alternansucrase to dextransucrase and to produce a high proportion of alternan to dextran, either fermentatively by cultivation of the strains on sucrose, or enzymatically by using alternansucrase produced by the strains of the invention. The selected strains can also be used to obtain microorganism strains having further improvements.

Growing community of inventors

Edwards, IL, United States of America

Gregory L Cote