Isocyclomaltooligosaccharide(s), isocyclomaltooligosaccharide-forming enzyme, their preparation and uses
A non-reducing saccharide by providing a novel non-reducing saccharide composed of glucose as constituents, a novel enzyme forming the non-reducing saccharide, a method and process for producing the same, a DNA encoding the enzyme, a recombinant DNA and transformant comprising the DNA, a composition comprising the non-reducing saccharide, and uses thereof are provided by use of an isocyclomaltooligosaccharide(s) having a structure represented by Formula 1, Cyclo{→6)-[α-D-Glcp-(1→4)] n -α-D-Glcp-(1→}, wherein “ n” is 4 or 5.
1. A purified isocyclomaltooligosaccharide-forming (ICM-forming) enzyme which has
an activity of forming an isocyclomaltooligosaccharide having a structure represented by formula 1, from α-1,4 glucan having a glucose polymerization degree of 3 or higher, wherein formula 1 is
Cyclo{→6)-[α-D-Glcp-(1→4)] n -α-D-Glcp-(1→}
and “n” is 4; and
said enzyme comprises an amino acid sequence identical to SEQ ID NO:2 or differing therefrom by a single amino acid deletion, replacement or addition.
2. The purified isocyclomaltooligosaccharide-forming enzyme of claim 1 wherein said α-1,4 glucan having a glucose polymerization degree of 3 or higher is one or more saccharides selected from the group consisting of maltooligosaccharide, maltodextrin, amylodextrin, amylose, amylopectin, soluble starch, liquefied starch, gelatinized starch, and glycogen.
3. The enzyme of claim 1 , which comprises an amino acid sequence identical to SEQ ID NO:2, or differing therefrom by a single amino acid replacement.
4. The enzyme of claim 1 which comprise an amino acid sequence identical to SEQ ID NO:2.
5. The enzyme of claim 1 whose amino acid sequence consists of the amino acid sequence of SEQ ID NO:2.
6. The enzyme of claim 1 which consists of the amino acid sequence of SEQ ID NO:2, or an amino acid sequence differing therefrom by a single amino acid deletion, replacement or addition.
7. The enzyme of claim 1 , having the following physicochemical properties:
(1) Molecular weight
About 106,000±20,000 daltons on SDS-gel electrophoresis;
(2) Isoelectric point
pI 7.5±0.5 on isoelectrofocusing a carrier ampholyte;
(3) Optimum temperature
50 to 55° C. when reacted at pH 6.0 for 30 min;
(4) Optimum pH
pH 4.5 to 8.0 when reacted at 30° C. for 30 min;
(5) Thermal stability
Stable up to the temperature of 35° C. when incubated at pH 6.0 for 60 min;
Stable up to the temperature of 40° C. when incubated at pH 6.0 for 60 min in the presence of 1 mM Ca 2+ ion; and
(6) pH Stability
Stable in a range of pH 4.5 to 9.0 when incubated at 4° C. for 24 hours.