*2.5. Formulation of the Holoenzyme Cocktail (HEC)*

The substrate specificities of the MFE-5E, MFE-5H, and MFE-45 enzymes were performed according to Mafa et al. [23], and the released total reducing sugars were measured according to a modified 3,5-dinitrosalicylic acid (DNS) method [24]. An amount of 1% (*w*/*v*) beechwood xylan, wheat arabinoxylan, Avicel, xyloglucan, locust bean gum and CMC were used as substrates. For *p*NP based substrate specificity assays, 2 mM *p*NP-A, *p*NP-G, *p*NP-M, *p*NP-C and *p*NP-X were used. Four GH enzymes; exo-glucanase (Exg-D), MFE-5E, MFE-5H and MFE-45, derived from the termite bacterial hindgut metagenome were used to formulate the holocellulolytic enzyme cocktail HEC-H (60% MFE-5H, 20% MFE-5E, 10% MFE-45 and 10% Exg-D). HEC-H was then supplemented with 10% protein loading of each of the following enzymes, *Aspergillus niger* β-glucosidase (Novozyme 188) and *Selenomonas ruminantium* xylosidase, SXA, during the hydrolysis of pretreated feedstocks. The reactions were initiated by adding the HEC-H and auxiliary enzymes to 2% (*w*/*v*) of untreated, Ca(OH)<sup>2</sup> and NaOH pretreated SSB or CC biomass samples suspended in 50 mM sodium citrate buffer at pH 5.5. The reaction was carried out by incubating the samples at 37 ◦C in an incubation room. The release of the total reducing sugars was measured using the DNS assays.
