TERNARY MIXTURES OF 6'-SL, LNnT and LST c

Abstract

A mixture of human milk oligosaccharides that consists essentially of 6′-SL, LNnT and LSTc, made by treating 6′-SL and LNnT in the presence of an α2,6-transsialidase.

1 . A mixture of human milk oligosaccharides (HMOs) consisting essentially of 6′-O-sialyllactose (6′-SL), lacto-N-neotetraose (LNnT) and sialyllacto-N-tetraose c (LST c). 
     
     
         2 . The mixture according to  claim 1 , wherein the molar ratio of LST c relative to the combination of 6′-SL and LNnT is at least 1:18. 
     
     
         3 . The mixture according to  claim 1  wherein the molar ratio of 6′ SL relative to LNnT is 0.18-5.5. 
     
     
         4 . A mixture of HMOs consisting essentially of 6′-O-sialyllactose lacto-N-neotetraose (LNnT), sialyllacto-N-tetraose c (LST c) and lactose. 
     
     
         5 . The mixture according to  claim 4 , wherein
 the molar ratio of the combination of 6′-SL and LNnT relative to LST c is 2-18, and   the molar ratio of lactose relative to LST c is about 1.   
     
     
         6 . The mixture according to  claim 4 , wherein one of the molar ratios 6′-SL to LST c and LNnT to LST c is not more than 4. 
     
     
         7 . The mixture according to  claim 4 , wherein the molar ratio of 6′-SL to LNnT is 0.18-5.5. 
     
     
         8 . The mixture according to  claim 1 , obtained by a method comprising the steps of reacting 6′-SL and LNnT in the presence of an α2,6-transsialidase to produce a reaction medium, and then removing lactose and the α2,6-transsialidase from the reaction medium. 
     
     
         9 . The mixture according to  claim 4 , obtained by a method comprising the steps of reacting 6′-SL and LNnT in the presence of an α2,6-transsialidase to produce a reaction medium, and then removing the α2,6-transsialidase from the reaction medium. 
     
     
         10 . The mixture according to  claim 9 , comprising the step of reacting 6′-SL and LNnT in a molar ratio of 1:3 to 3:1 in the presence of an α2,6-transsialidase having a conversion rate of at least 20%, up to about 55% for the reaction of 6′-SL and LNnT. 
     
     
         11 . The mixture according to  claim 10 , comprising the step of reacting 6′-SL and LNnT in a molar ratio of 1:1, in the presence of an α2,6-transsialidase having a conversion rate of at least 20%, up to about 45%, for the reaction of 6′-SL and LNnT. 
     
     
         12 . The mixture according to  claim 10 , wherein the conversion rate for the reaction of 6′-SL and LNnT is 30-45%. 
     
     
         13 . The mixture according to  claim 8 , wherein the α2,6-transsialidase is a mutated α2,6-transsialidase that comprises has an amino acid sequence with at least 60% identity to SEQ ID No. 1, and further comprises at least one mutation selected from the group consisting of:
 at position 156, an amino acid selected from the group consisting of Ser, Thr, Cys, Tyr, Asn, Gln and Trp, 
 at position 161, an amino acid selected from the group consisting of Ala, Val, Ile, Leu, Phe, Tyr, Trp and Gly, 
 at position 180, an amino acid selected from the group consisting of Asp, Asn and Gln; 
 at position 186, an amino acid selected from the group consisting of Val, Ile, Leu, Met, Phe, Tyr, Trp, Ser, Cys and Thr; 
 at position 218, an amino acid selected from the group consisting of Val, Ile, Leu, Met, Phe, Tyr, Trp, Cys, Gly and Thr; 
 at position 222, an amino acid selected from the group consisting of Gln, Asp, Glu, Cys, Thr, Phe, Tyr, Trp, Arg, Lys and His; 
 at position 235, an amino acid selected from the group consisting of Arg, His, Ser, Cys, Ala, Val, Ile or Leu; 
 at position 242, an amino acid selected from the group consisting of Arg, His and Lys; 
 at position 261, an amino acid selected from the group consisting of His, Lys, Asp, Glu, Ala, Val, Leu and Phe; 
 at position 315, an amino acid selected from the group consisting of Ser, Thr and Cys; 
 at position 342, an amino acid selected from the group consisting of Ser and Cys; 
 at position 349, an amino acid selected from the group consisting of Ser, Thr and Cys; 
 at position 350, an amino acid selected from the group consisting of Ser, Thr, Cys, Tyr, Trp and Phe; 
 at position 356, an amino acid selected from the group consisting of Ala, Val, Ile, Leu, Phe and Trp; and 
 at position 438, an amino acid selected from Arg, His and Lys. 
 
     
     
         14 . The mixture according to  claim 13 , wherein the at least one mutation is selected from the group consisting of:
 at position 156 Gly is substituted by Ser, Cys or Tyr;   at position 161 Gln is substituted by Phe or Gly;   at position 180 Glu is substituted by Asp,   at position 186 Ala is substituted by Tyr, Cys or Leu;   at position 218 Ala is substituted by Ile, Val, Phe or Tyr;   at position 222 Asn is substituted by Cys, Asp, Arg or Phe;   at position 235 Lys is substituted by Arg, His, Cys or Val;   at position 242 Val is substituted by His;   at position 261 Arg is substituted by Asp, Phe, His or Val;   at position 315 Leu is substituted by Cys;   at position 342 Thr is substituted by Cys;   at position 349 Gly is substituted by Ser or Cys;   at position 350 Gly is substituted by Ser, Tyr, Phe or Cys;   at position 356 Tyr is substituted by Val or Phe; and   at position 438 Pro is substituted by His.   
     
     
         15 . The mixture according to  claim 14 , wherein the mutated α2,6-transsialidase has a mutation at least at two amino acid positions from 156, 218, 222 and 349. 
     
     
         16 . An anti-infective composition for treating bacterial infections comprising the mixture of  claim 1 . 
     
     
         17 .- 18 . (canceled) 
     
     
         19 . An anti-infective composition for treating bacterial infections comprising the mixture of  claim 4 . 
     
     
         20 . An anti-infective composition for treating bacterial infections comprising the mixture of  claim 8 . 
     
     
         21 . An anti-infective composition for treating bacterial infections comprising the mixture of  claim 9 .