## 4 Discussion
### Secretion of Enzymes Related to Degradation of Lignocellulose
It was previously thought that oxidative enzymes were only involved in lignin degradation. Recently, however, the degradation of cellulose and hemicellulose, in which LPMOs play an essential role, has been actively studied [62]. Today, LPMOs are grouped into eight families of CAZymes (AA9-AA11 and AA13-AA17) and have been shown to oxidatively cleave various oligosaccharidesarides (such as cellulose, hemicellulose, pectin, starch and chitin) [63]. A more in-depth study of LPMO sequences found in the _C. laeev_ secretomes revealed that they contain the DUF6595 domain (pfam20238), similarly to recently characterized members of a new LPMO family from _Laetisaria arvalis_ (defined as X325), which represents a widespread family of copper-containing proteins in various fungal genomes [64]. Although the overall structural fold of the X325 proteins was similar to that of the LPMOs, they did not possess activity against the polysaccharides tested and their functions may vary in different fungi. Despite the typical conservative pattern of X325 proteins, the identity between LPMOs from _C. laeev_ secretomes and X325 LPMO from _L. arvalis_ (MK088083) is only about 30% (Supplementary Figure S2). It is worth noting that the absence of enzymatic activity of various LPMOsis quite common under model experimental conditions. In work [65], two AA14 LPMOs (_PcAA_14A and _PcAA_14B) from the WRF _Pycnoporus occineus_ were characterized to be inactive against model substrates such as wheat arabinxylan, arabinar, barley \(\beta\)-glucan, curdlan, konjac glucomannan, tamarind xyloglucan, starch, Avicel, chitin, laminin, PASC and birch xylan. However, pine and poplar substrates pretreated with the _Trichoderma reesei_ cellulase-xylanase cocktail were associated with an increase in saccharification ability with the addition of _Pc_AA14A and _Pc_AA14B LPMOs. It has also been shown that different LPMO isoenzymes can have different preferences for different types of oligosaccharides [66]. These studies, which indicate a close relationship between the chemical structure of plant polysaccharides and their interaction with the substrate-binding surface of LPMOs, shed light on a wide variety of LPMO genes in fungal genomes. In addition to LPMOs, loosenin-like expansins were found in the secretomes of _C. laeve_. Moreover, increased secretion was showed for some of them on the sawdust-containing media, and also new proteins were detected. Expansin-like proteins can modify the surface of various lignocellulosic substrates in a non-enzymatic way, thus enhancing the enzymatic degradation of lignocellulosic [67; 68]. The synergistic effect of expansins and LPMOs has also been reported [69]. In addition, the combination of xylanase with _Cmi_EXLX2 and _Daq_EXLX1 expansins increased product yield from hardwood pulp by approximately 25%, while _Tr_AA9A LPMO from _Trichoderma reesei_ supplemented with the same expansins increased total product yield by over 35%. A recent study demonstrated the differences in the specificity of the interaction of different expansins with various polysaccharides. It was suggested that the multiplicity of expansins in an organism and the fine regulation of their expression may in part be caused by the specific binding of individual expansins to particular cell wall polysaccharides [70]. This is in agreement with the differential secretion of the _C. laeve_ loosenin-like expansins on media containing different types of sawdust detected in our study (Figure 4). In addition to the specific molecular structure of the polysaccharides, LPMO activity also depends on the presence of an electron donor (i.e., the so-called reducing agent) in a reaction medium. Small organic molecules (such as ascorbic acid, cysteine, glutathione, gallic acid, etc.) synthesized by the fungus or lignin-derived during lignocellulosic destruction, as well as enzymatic redox partners (flavo-enzymes AA3 and AA7 family CAZymes) can act as reducing agents for different LPMOs [71; 72].