3.1 Ganoderic acids biosynthesis pathway
Ganoderic acids(GAs)is a kind of triterpenoids and the main medicinal component of G.lucidum with function of anti-tumor，immuno-reglulation，and anti-oxidant et al.［8］.Currently，most studies about triterpenoid biosynthesis were in plants，while the detailed biosynthesis pathways in fungi were studied much less.Shiao［27］proved that GAs were synthesized via MVA pathway by using isotopic tracer experiments.In this study，we found that the genes only existed in the MVA pathway of terpenoids backbone biosynthesis in G.lucidum genome while no genes in the MEP pathway，which is another terpenoids biosynthesis pathway in plants.Thus，at the genome level，we confirmed the terpenoids biosynthesis in G.lucidum and even in fungi was only through the MVA subway，not the MEP pathway.Interestingly，we identified a fusion gene(metC-MVK)in basidiomycetes，half of which was homologous to the MVK enzyme in terpenoids biosynthesis and half of which was similar to the metC enzyme，a cystathionine beta-lyase.In animal and even Ascomycota fungi，they are two separate genes，suggesting the appearance of metC-MVK occurred in the ancestor of Basidiomycota fungi.It is unknown why the two genes fused and whether the fusion gene has the function of both separate genes.
We have characterized all the enzymes in the terpenoid backbone biosynthesis in G.lucidum(Table 5).In triterpenoid synthesis，2，3-epoxysqualene is the precursor and the difference of cyclization，oxidation，hydroxylation and glycosylation leads to form different triterpenoids［28］.However，there is little knowledge about the pathway that lies downstream of cyclization.It has been reported that these modification are carried out by cytochrome P450，glycosyltransferases and other enzymes［28］.P450 are speculated to be involved in a wide range of modification，including oxidation and hydroxylation in the synthesis of triterpenoids［29］.Due to the large number and diversity，it is difficult to identify their specific functions based on homology.There are only four P450 genes reported to involve in the biosynthesis of triterpenoid saponins［29- 31］.So far no P450 genes cloned in GAs biosynthesis.In G.lucidum，we characterized 222 genes encoding P450 enzymes and 21 of them were very similar with the four known P450 genes.The plentiful of putative P450 genes provided the potential of different oxidation and hydroxylation，thus plentiful GAs formed in G.lucidum.(https://www.daowen.com)
Glycosylation，transferring the active saccharides to the triterpenoid backbones，stabilizing the product and altering its physiological activity［32］，is the last and key modification in GAs biosynthesis.UGTs are reported to contribute the glycosylation in triterpenoid biosynthesis and so far only six UGTs are experimentally identified in the triterpenoid biosynthesis［30，31］.Using the known UGTs sequences［29］to search against G.lucidum genes，we found six putative UGTs in G.lucidum genome，which might be responsible for the glycosylation modification in the GAs biosynthesis in G.lucidum.Among these six putative UGTs，the gene(G_lucidum_10009504)was highly similar with UGT73K1，which glycosylated both triterpenoids and(iso)flavones in Medicago truncatula［33］.Using the known UGTs sequences to search other fungi，we found that there were several UGT homologs in the Agaricomycotina fungi but none of them in the Ascomycota fungi.This might suggest that the ascomycetes use other glycosylation enzymes not these UGTs in the triterpenoid synthesis.