Part:BBa_K4382005

Arabinofuranoside - XynD

Usage

When Xylanase D was characterized, it was first mischaracterised as an endoxylanase - due to a great deal of sequence homology seen with previously identified endoxylanases. However, further functional analysis performed by Bourgois et. al. in 2017 showed that the designated substrate for XynD is arabinoxylo-oligosaccharides(AXOS) - wherein it cleaves the bond between the xylan polymer and the arabinose side chain. In our project, we are cleaving the side chains.

Biology

XynD obtained from Bacillus subtilis 168 belongs to the GH-43 family of glycosyl hydrolases, with a Family 6 Carbohydrate Binding Module at the C terminal. The enzyme works optimally at 450C and a pH of 5.6.

In terms of substrate preference, it was observed that XynD acts only on specifically substituted Arabinoxylans. It was found that only C-(O)-2 and C-(O)-3 arabinoses were removed by the xynD - with 80 percent efficiency. This value is independent of the degree of substitution of the xylan backbone (as shown below through a common decrease in the A/X ratio). One exception to this process is wheat bran AXOS, where A/X is 0.23 and only 41% of the substituents were acted upon by the enzyme.

An interesting observation is that in the Bacillus subtilis 168 genome, it is seen that XynD is present downstream of the same promoter as XynC, which indicates some synergy in their activities.

References

1. Bourgois, T. M., Van Craeyveld, V., Van Campenhout, S., Courtin, C. M., Delcour, J. A., Robben, J., & Volckaert, G. (2007, April 11). Recombinant expression and characterization of XynD from Bacillus subtilis subsp. subtilis ATCC 6051: a GH 43 arabinoxylan arabinofuranohydrolase. Applied Microbiology and Biotechnology, 75(6), 1309–1317. https://doi.org/10.1007/s00253-007-0956-2

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal EcoRI site found at 111
  Illegal SpeI site found at 82
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 111
  Illegal SpeI site found at 82
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal EcoRI site found at 111
  Illegal BamHI site found at 147
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal EcoRI site found at 111
  Illegal SpeI site found at 82
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal EcoRI site found at 111
  Illegal SpeI site found at 82
  Illegal NgoMIV site found at 610
  Illegal NgoMIV site found at 1024
  Illegal AgeI site found at 595
  Illegal AgeI site found at 1453
  
• 1000
  COMPATIBLE WITH RFC[1000]