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Biotechnol Biofuels. 2016 Jul 8;9:140. doi: 10.1186/s13068-016-0550-x. eCollection 2016.

The GH51 α-l-arabinofuranosidase from Paenibacillus sp. THS1 is multifunctional, hydrolyzing main-chain and side-chain glycosidic bonds in heteroxylans.

Author information

1
UBMB, Université de Tunis El Manar, BP 94, 1068 Rommana, Tunisia.
2
Laboratoire des Ressources Sylvo-Pastorales, Institut Sylvo-Pastoral de Tabarka, Institution de la Recherche et de l'Enseignement Supérieur Agricoles, Université de Jendouba, Jendouba, Tunisia.
3
CNRS, INRA, INSA, LISBP, Université de Toulouse, Toulouse, France.
4
Institute of Biological, Environmental and Rural Sciences, Aberystwyth University, Aberystwyth, SY23 3DA Ceredigion UK.
5
INRA, FARE, Université de Reims Champagne Ardenne, 2, Esplanade Roland Garros, 51100 Reims, France.
#
Contributed equally

Abstract

BACKGROUND:

Conceptually, multi-functional enzymes are attractive because in the case of complex polymer hydrolysis having two or more activities defined by a single enzyme offers the possibility of synergy and reduced enzyme cocktail complexity. Nevertheless, multi-functional enzymes are quite rare and are generally multi-domain assemblies with each activity being defined by a separate protein module. However, a recent report described a GH51 arabinofuranosidase from Alicyclobacillus sp. A4 that displays both α-l-arabinofuranosidase and β-d-xylanase activities, which are defined by a single active site. Following on from this, we describe in detail another multi-functional GH51 arabinofuranosidase and discuss the molecular basis of multifunctionality.

RESULTS:

THSAbf is a GH51 α-l-arabinofuranosidase. Characterization revealed that THSAbf is active up to 75 °C, stable at 60 °C and active over a broad pH range (4-7). THSAbf preferentially releases para-nitrophenyl from the l-arabinofuranoside (k cat/K M = 1050 s(-1) mM(-1)) and to some extent from d-galactofuranoside and d-xyloside. THSAbf is active on 4-O-methylglucuronoxylans from birch and beechwood (10.8 and 14.4 U mg(-1), respectively) and on sugar beet branched and linear arabinans (1.1 ± 0.24 and 1.8 ± 0.1 U mg(-1)). Further investigation revealed that like the Alicyclobacillus sp. A4 α-l-arabinofuranosidase, THSAbf also displays endo-xylanase activity, cleaving β-1,4 bonds in heteroxylans. The optimum pH for THASAbf activity is substrate dependent, but ablation of the catalytic nucleophile caused a general loss of activity, indicating the involvement of a single active center. Combining the α-l-arabinofuranosidase with a GH11 endoxylanase did not procure synergy. The molecular modeling of THSAbf revealed a wide active site cleft and clues to explain multi-functionality.

CONCLUSION:

The discovery of single active site, multifunctional enzymes such as THSAbf opens up exciting avenues for enzyme engineering and the development of new biomass-degrading cocktails that could considerably reduce enzyme production costs.

KEYWORDS:

Biomass; Enzyme cocktails; Glycoside hydrolase; Wheat bran; Xylanase

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