Cellulose induced protein 1 (Cip1) from Trichoderma reesei enhances the enzymatic hydrolysis of pretreated lignocellulose

Abstract

Background: Trichoderma reesei is currently the main strain for the commercial production of cellulase. Cellulose induced protein 1 (Cip1) is one of the most abundant proteins in extracellular proteins of T. reesei. Reported literatures about Cip1 mainly focused on the regulation of Cip1 and its possible enzyme activities, but the effect of Cip1 on the enzymatic hydrolysis of lignocellulose and possible mechanism have not still been reported.

Results: In this study, Cip1 from T. reesei was cloned, expressed and purified, and its effects on enzymatic hydrolysis of several different pretreated lignocellulose were investigated. It was found that Cip1 could promote the enzymatic hydrolysis of pretreated lignocellulose, and the promoting effect was significantly better than that of bovine serum albumin (BSA). And especially for the lignocellulosic substrate with high lignin content such as liquid hot water pretreated corn stover and corncob residue, the promoting effect of Cip1 was even better than that of the commercial cellulase when adding equal amount protein. It was also showed that the metal ions Zn²⁺ and Cu²⁺ influenced the promoting effect on enzymatic hydrolysis. The Cip1 protein had no lyase activity, but it could destroy the crystal structure of cellulose and reduce the non-productive adsorption of cellulase on lignin, which partly interpreted the promoting effect of Cip1 on enzymatic hydrolysis of lignocellulose.

Conclusion: The Cip1 from T. reesei could significantly promote the enzymatic hydrolysis of pretreated lignocellulose, and the promotion of Cip1 was even higher than that of commercial cellulase in the enzymatic hydrolysis of the substrates with high lignin content. This study will help us to better optimize cellulase to improve its ability to degrade lignocellulose, thereby reducing the cost of enzymes required for enzymatic hydrolysis.

Keywords: Cellulase; Cellulose induced protein 1; Crystal structure; Enzymatic hydrolysis; Lignocellulose.

Fig. 1

SDS-PAGE analysis of Cip1. Lane M: protein marker; lane 1: purified Cip1; lane 2 and 3: different concentrations of Cip1 after Endo H treatment

Fig. 2

Effect of BSA, commercial cellulase SP and Cip1 addition on enzymatic hydrolysis of CS (a), LPCS (b), NPCS (c), CC (d), CCR (e) and Avicel (f) at 24 h, 48 h and 72 h. In which, protein dosage of 0.5 mg/g DM was added. The data marked with ** represented there were significant difference (p < 0.05) and an asterisk * means that the difference was not significant (p > 0.05)

Fig. 3

The effect of metal ions and EDTA addition on the promotion of Cip1. No metal ions and chemical reagents were added as the control, and its relative glucan conversion was defined as 100%. The final concentration of metal ions and EDTA was 4 mM in hydrolysis system. The data marked with ** and * respectively means there was significant difference (p < 0.05) and not significant difference (p > 0.05)

Fig. 4

XRD spectra of solid residues after 72 h of enzymatic hydrolysis of CCR by cellulase or buffer with and without the addition of Cip1 and Cip1-CD

Fig. 5

a The relative filter paper activity in the supernatant for hydrolysis of CCR at 72 h. b The relative activities of FPA, CBH, EG and BG in the supernatant after lignin adsorption for 24 h. The data marked with ** and * respectively means there was significant difference (p < 0.05) and not significant difference (p > 0.05). The relative activity at 0 h of reaction time was defined as 100%

Fig. 6

Predicted model of Cip1 enhanced enzymatic hydrolysis of lignocellulose with cellulase