Development of an ultra-high-temperature process for the enzymatic hydrolysis of lactose. I. The properties of two thermostable beta-glycosidases.

Recombinant beta-glycosidases from hyperthermophilic Sulfolobus solfataricus (SsbetaGly) and Pyrococcus furiosus (CelB) have been characterized with regard to their potential use in lactose hydrolysis at about 70 degrees C or greater. Compared with SsbetaGly, CelB is approximately 15 times more stable against irreversible denaturation by heat, its operational half-life time at 80 degrees C and pH 5.5 being 22 days. The stability of CelB but not that of SsbetaGly is decreased 4-fold in the presence of 200 mM lactose at 80 degrees C. CelB displays a broader pH/activity profile than SsbetaGly, retaining at least 60% enzyme activity between pH 4 and 7. Both enzymes have a similar activation energy for lactose hydrolysis of approximately 75 kJ/mol (pH 5.5), and this is constant between 30 and 95 degrees C. D-Galactose is a weak competitive inhibitor against the release of D-glucose from lactose (Ki approximately 0.3 M), and at 80 degrees C the ratio of Ki, D-galactose to Km,lactose is 2.5 and 4.0 for CelB and SsbetaGly, respectively. SsbetaGly is activated up to 2-fold in the presence of D-glucose with respect to the maximum rate of glycosidic bond cleavage, measured with o-nitrophenyl beta-D-galactoside as the substrate. By contrast, CelB is competitively inhibited by D-glucose and has a Ki of 76 mM. The transfer of the galactosyl group from lactose to acceptors such as lactose or D-glucose rather than water is significant for both enzymes and depends on the initial lactose concentration as well as the time-dependent substrate/product ratio during batchwise lactose conversion. It is approximately 1.8 times higher for SsbetaGly, compared with CelB. Overall, CelB and SsbetaGly share their catalytic properties with much less thermostable beta-glycosidases and thus seem very suitable for lactose hydrolysis at >/=70 degrees C.

[1]  L. Pearl,et al.  Crystal structure of the beta-glycosidase from the hyperthermophilic archeon Sulfolobus solfataricus: resilience as a key factor in thermostability. , 1997, Journal of molecular biology.

[2]  R. Crittenden,et al.  Production, properties and applications of food-grade oligosaccharides , 1996 .

[3]  R. Furneaux,et al.  Use of Enzyme Technology to Convert Waste Lactose into Valuable Products , 1996 .

[4]  M. Siso.  The biotechnological utilization of cheese whey: A review , 1996 .

[5]  B. Saha,et al.  Thermostable β-Glucosidases , 1996 .

[6]  R. Furneaux,et al.  Synthesis of allyl β-d-galactopyranoside from lactose using Streptococcus thermophilus β-d-galactosidase , 1996 .

[7]  L. Fischer,et al.  Catalytical Potency of β-Glucosidase from the Extremophile Pyrococcus furiosus in Glucoconjugate Synthesis , 1996, Bio/Technology.

[8]  W. D. de Vos,et al.  Characterization of the celB gene coding for beta-glucosidase from the hyperthermophilic archaeon Pyrococcus furiosus and its expression and site-directed mutation in Escherichia coli , 1995, Journal of bacteriology.

[9]  M. Rossi,et al.  Expression and extensive characterization of a beta-glycosidase from the extreme thermoacidophilic archaeon Sulfolobus solfataricus in Escherichia coli: authenticity of the recombinant enzyme. , 1995, Enzyme and microbial technology.

[10]  S. d'Auria,et al.  A thermostable β-glycosidase from Sulfolobus solfataricus: temperature and SDS effects on its functional and structural properties , 1995 .

[11]  L. Pearl,et al.  Thermostable β-glycosidase from Sulfolobus solfataricus , 1994 .

[12]  A. Stams,et al.  An extremely thermostable β-glucosidase from the hyperthermophilic archaeon Pyrococcus furiosus : a comparison with other glycosidases , 1994 .

[13]  H. Tomomatsu.  Health effects of oligosaccharides , 1994 .

[14]  A. Stams,et al.  Purification and characterization of an extremely thermostable beta-glucosidase from the hyperthermophilic archaeon Pyrococcus furiosus. , 1993, European journal of biochemistry.

[15]  M. Rossi,et al.  Exo‐glucosidase activity and substrate specificity of the beta‐glycosidase isolated from the extreme thermophile Sulfolobus solfataricus , 1993, Biotechnology and applied biochemistry.

[16]  S. Honda,et al.  Two-dimensional mapping of N-glycosidically linked asialo-oligosaccharides from glycoproteins as reductively pyridylaminated derivatives using dual separation modes of high-performance capillary electrophoresis. , 1992, Analytical biochemistry.

[17]  C. H. Amundson,et al.  Hydrolysis of lactose in skim milk by immobilized β‐galactosidase (bacillus circulans) , 1992, Biotechnology and bioengineering.

[18]  D. Grogan.  Evidence that β-Galactosidase of Sulfolobus solfataricus Is Only One of Several Activities of a Thermostable β-d-Glycosidase , 1991 .

[19]  C. H. Amundson,et al.  Use of novel immobilized β-galactosidase reactor to hydrolyze the lactose constituent of skim milk , 1991 .

[20]  M. Roeckel,et al.  Production of Thermostable β‐Galactosidase with Thermoanaerobacter ethanolicusa , 1990 .

[21]  M. Sinnott,et al.  Catalytic mechanism of enzymic glycosyl transfer , 1990 .

[22]  C. H. Amundson,et al.  Use of novel immobilized β-galactosidase reactor to hydrolyze the lactose constituent of skim milk , 1990 .

[23]  R. E. Huber,et al.  Determination of the roles of Glu-461 in beta-galactosidase (Escherichia coli) using site-specific mutagenesis. , 1990, The Journal of biological chemistry.

[24]  S. Zárate,et al.  Oligosaccharide Formation During Enzymatic Lactose Hydrolysis: A Literature Review. , 1990, Journal of food protection.

[25]  R. Rella,et al.  Thermostable β-galactosidase from the archaebacterium Sulfolobus solfataricus Purification and properties , 1990 .

[26]  R. S. Peterson,et al.  Lactose hydrolysis by immobilized β‐galactosidase in capillary bed reactor , 1989 .

[27]  C. H. Amundson,et al.  Hydrolysis of lactose in skim milk by immobilized β‐galactosidase in a spiral flow reactor , 1989, Biotechnology and Bioengineering.

[28]  B. Chang,et al.  Purification and thermostability of β-galactosidase (lactase) from an autolytic strain of Streptococcus salivarius subsp. thermophilus , 1989, Journal of Dairy Research.

[29]  N. Scrimshaw,et al.  The acceptability of milk and milk products in populations with a high prevalence of lactose intolerance. , 1988, The American journal of clinical nutrition.

[30]  J. Bourne,et al.  Formation of oligosaccharides during enzymatic lactose hydrolysis and their importance in a whey hydrolysis process: Part II: Experimental , 1987, Biotechnology and bioengineering.

[31]  J. Bourne,et al.  Formation of oligosaccharides during enzymatic lactose: Part I: State of art , 1987, Biotechnology and bioengineering.

[32]  R. Daniel,et al.  Purification and properties of a stable beta-glucosidase from an extremely thermophilic anaerobic bacterium. , 1987, The Biochemical journal.

[33]  V. Gekas,et al.  Hydrolysis of lactose: a literature review , 1985 .

[34]  R. R. Mahoney,et al.  Modification of lactose and lactose-containing dairy products with β-galactosidase. , 1985 .

[35]  D. Cowan,et al.  Some properties of a β‐galactosidase from an extremely thermophilic bacterium , 1984 .

[36]  A. Renken,et al.  The kinetic of lactose hydrolysis for the β‐galactosidase from Aspergillus niger , 1982 .

[37]  M. Sinnott,et al.  The effect of methanol and dioxan on the rates of the beta-galactosidase-catalysed hydrolyses of some beta-D-galactrophyranosides: rate-limiting degalactosylation. The ph-dependence of galactosylation and degalactosylation. , 1973, The Biochemical journal.