Structure and properties of the extracellular inulinase of Kluyveromyces marxianus CBS 6556

Abstract

In the yeast Kluyveromyces marxianus two forms of inulinase were present, namely, an inulinase secreted into the culture fluid and an inulinase retained in the cell wall. Both forms were purified and analyzed by denaturing and nondenaturing polyacrylamide gel electrophoresis. With the use of endo-ID-N-acetyl-glucosaminidase H, it was established that the enzyme retained in the cell wall and the enzyme secreted into the culture fluid have similar subunits consisting of a 64-kDa polypeptide with varying amounts of carbohydrate (26 to 37% of the molecular mass). The two forms of inulinase differed in size because of their differences in subunit aggregation. The enzyme present in the culture fluid was a dimer, and the enzyme retained in the cell wall was a tetramer. The differences in oligomerization did not affect the apparent Km values towards the substrates sucrose and raffinose. These findings support the hypothesis that the retention of glycoproteins in the yeast cell wall may be caused by a permeability barrier towards larger glycoproteins. The amino-terminal end of inulinase was determined and compared with the amino terminus of the closely related invertase. The kinetic and structural evidence indicates that in yeasts two distinct ,3-fructosidases exist, namely, invertase and inulinase.

Inactivation and conformational changes of yeast invertase during unfolding in urea and guanidinium chloride solutions

“…However, the results clearly show that inactivation occurs before noticeable conformational change of the enzyme molecule as a whole can be detected. 8). It appears that the Trp residues are partly situated on the surface of the enzyme molecule and partly buried in its interior, and the latter are not easily exposed by low concentrations of urea or guanidinium chloride.…”

Biochemical and molecular identification and characterization of lactic acid bacteria and yeasts isolated from Ethiopian naturally fermented buttermilk

“…Capacity of this yeast to utilize lactose indicates its involvement in fermentation of SFB (Ramirez-Zavala et al 2004). The yeast is utilized in the synthesis of commercial enzymes including: Inulinase production of fructose syrup (Rouwenhorst et al 1990); β -galactosidase, reduction of lactose (Martins et al 2002); protein phosphatase, modification of cheese making qualities of casein and carboxypeptidases (reduction of bitter taste) and aminopeptidases (direct processing and ageing of dairy and meat products . This yeast has high metabolic diversity and is used for different industrial application (Lane and Morrissey 2010).…”

Purification and properties of an extracellular inulinase-like β-fructosidase from Bacillus stearothermophilus

“…1987). A 350-450 kDa inulinase has also been identified in KIuyverom,yccs marxianus (Rouwenhorst et al 1990b) which was composed of 64 kDa subunits.…”