A novel chimaeric flocculation protein enhances flocculation in .

Yeast flocculation is the reversible formation of multicellular complexes mediated by lectin-like cell wall proteins binding to neighbouring cells. Strong flocculation can improve the inhibitor tolerance and fermentation performance of yeast cells in second generation bioethanol production. The strength of flocculation increases with the size of the flocculation protein and is strain dependent. However, the large number of internal repeats in the sequence of from S288c makes it difficult to recombinantly express the gene to its full length. In the search for novel flocculation genes resulting in strong flocculation, we discovered a DNA sequence, , that gives NewFlo phenotype flocculation in CEN.PK 113-7D. The nucleotide sequence of the internal repeats of differed from those of . We hypothesized that a chimaeric flocculation gene made up of a variant derived from S288c and additional repeats from from CCUG 53310 would be more stable and easier to amplify by PCR. The constructed gene, , had 22 internal repeats compared to 18 in . Expression of in otherwise non-flocculating strains led to strong flocculation. Despite the length of the gene, the cassette containing could be easily amplified and transformed into yeast strains of different genetic background, leading to strong flocculation in all cases tested. The developed gene can be used as a self-immobilization technique or to obtain rapidly sedimenting cells for application in sequential batches without need for centrifugation.