An Insight into the Essential Role of Carbohydrate-Binding Modules in Enzymolysis of Xanthan

Abstract

To date, due to the low accessibility of enzymes to xanthan substrates, the enzymolysis of xanthan remains deficient, which hinders the industrial production of functional oligoxanthan. To enhance the enzymatic affinity against xanthan, the essential role of two carbohydrate binding modules—MiCBMx and PspCBM84, respectively, derived from Microbacterium sp. XT11 and Paenibacillus sp. 62047—in catalytic properties of endotype xanthanase MiXen were investigated for the first time. Basic characterizations and kinetic parameters of different recombinants revealed that, compared with MiCBMx, PspCBM84 dramatically increased the thermostability of endotype xanthanase, and endowed the enzyme with higher substrate affinity and catalytic efficiency. Notably, the activity of endotype xanthanase was increased by 16 times after being fused with PspCBM84. In addition, the presence of both CBMs obviously enabled endotype xanthanase to produce more oligoxanthan, and xanthan digests prepared by MiXen-CBM84 showed better antioxidant activity due to the higher content of active oligosaccharides. The results of this work lay a foundation for the rational design of endotype xanthanase and the industrial production of oligoxanthan in the future.

Identification of a novel xanthan-binding module of a multi-modular Cohnella sp. xanthanase

“…The binding affinity of Cs CBM66 for XLT-xanthan adds a novel binding specificity to the CBM66 family. These findings have potential applications of enzyme modification for xanthan, as combinations of the binding module Cs CBM66 and catalytic modules for the cellulose-like backbone from GH5, GH9 or other glycoside hydrolase families could lead to the design of highly efficient enzymes ( Ni et al, 2023 ).…”

“…Generally, CBMs are thought to have various functions which vary depending on the individual CBMs and their cognate substrates. Such functions include the increase of enzyme concentration around the saccharide substrates ( Zverlov et al, 2001 ; Ni et al, 2023 ), the binding to specific regions of substrates ( Boraston et al, 2001 ; Cuskin et al, 2012 ), the disruption of the structure of substrates ( Giardina et al, 2001 ) and the adhesion to the surface of substrates ( Singh et al, 2014 ). CBMs play important roles in biocatalytic polymer decomposition processes.…”

Xanthan: enzymatic degradation and novel perspectives of applications

“…However, Mi Xen fused with Psp XBM showed a higher affinity for the substrate than Mi Xen equipped with its native XBM. Furthermore, the hybrid xanthanase has increased thermostability and greater affinity for the substrate, leading to enhanced enzymatic efficiency (Ni et al 2023 ). The use of XBMs for engineering xanthanases with altered or improved characteristics may find applications in the development of technologies for producing functional XTOS, such as oligosaccharides with antioxidant activity, as proposed by Ni et al ( 2023 ).…”