Isolation and characterization of the HFBIII hydrophobin of Trichoderma reesei

Hydrophobins are surface active proteins produced by filamentous fungi. They have a role in fungal growth as structural components and in the interaction of fungi with their environment. Due to their physico-chemical characteristics they are very interesting proteins and suited also for various applications. We have been characterizing the hydrophobins of T. reesei on molecular and functional level. Previously we have reported characterization of the HFBI and HFBII hydrophobins. Recently, we have isolated a third hydrophobin, HFBIII, from T. reesei. The hfb3 gene was cloned based on internal peptide sequences obtained for HFBIII found in the supernatant of lactose cultures. HFBIII is a classII hydrophobin and very similar to both HFBI and HFBII the most specific feature of it being an extra cysteine residue between the regular 4th and 5th cysteines. Expression of hfb3 was studied and revealed that it is expressed similar to hfb2 on complex plat materials and lactose but the expression levels are lower. To characterize HFBIII at a protein level, a T. reesei strain overexpressing HFBIII in a HFBII deletion background was constructed. A purification prodedure for HFBIII was developed. HFBIII is largely retained in the mycelium in vegetative cultures. Our findings show that different treatments are needed for removal of HFBIII from the mycelium than for HFBI, which can be found in the cell walls of glucose-grown mycelia. This can indicate different structural roles for the two proteins.