The Fps1p aquaglyceroporin facilitates the use of small aliphatic amides as a nitrogen source by amidase-expressing yeasts

Abstract

Saccharomyces cerevisiae acquires a resistance to high, toxic levels of acetic acid by destabilizing Fps1p, the plasma membrane aquaglyceroporin through which this acid - in its undissociated state - enters the cell. In this study, Fps1p loss was shown to confer resistances to acetic acid, acrolein and allyl alcohol, not just in S. cerevisiae but also in the osmotolerant spoilage yeast Zygosaccharomyces rouxii. However, in Z. rouxii, the loss of Fps1p severely compromised the use of acetamide and several other small amides as sources of nitrogen, an indication that these amides enter the cells of this yeast by passive diffusion through the Fps1p pore. Saccharomyces cerevisiae cannot grow on acetamide, but was conferred with an ability to use this and other small amides as nitrogen sources by heterologous expression of a Z. rouxii ORF (ZrAMD1) with protein sequence identity to the amdS-encoded amidase of Aspergillus nidulans. This capacity of ZrAMD1-expressing S. cerevisiae to assimilate amide nitrogen was severely compromised by the loss of Fps1p. ZrAMD1 appears to encode the major amidase of Z. rouxii as a Zramd1Delta deletant mutant had, like the Zrfps1Delta deletant, lost the ability to assimilate small amides as sources of nitrogen.

Comparative Polygenic Analysis of Maximal Ethanol Accumulation Capacity and Tolerance to High Ethanol Levels of Cell Proliferation in Yeast

“…The selection marker utilized was the amidase gene ( AMD1 ), which was amplified from the vector pF6a-AMD1-MX6. The gene AMD1 was cloned from Z. rouxii [49]. The primers utilized in the AMD1 amplification had at least 80 extra bases that corresponded to the flanking regions of the area to be deleted.…”

QTL Analysis of High Thermotolerance with Superior and Downgraded Parental Yeast Strains Reveals New Minor QTLs and Converges on Novel Causative Alleles Involved in RNA Processing

“…For the first transformation, a linear DNA fragment with the AMD1 gene from Zygosaccharomyces rouxii flanked by 50 bp sequences that are homologous to the two sides of the MKT1 ORF was amplified from plasmid pFA6a-AMD1-MX6 [48] by PCR, and transformed into 21A. Transformants were grown on YCB (Yeast Carbon Base 1.17%, phosphate buffer 3%, Bacto agar 2%) plates containing 10 mM acetamide.…”

Acrolein-stressed threshold adaptation alters the molecular and metabolic bases of an engineered Saccharomyces cerevisiae to improve glutathione production

“…This inspired us to explore a problematic molecular mechanism eliminating the beneficial fitness. To date, there is only one literature report that Acr might cross the cell membrane by means of aquaglyceroporin-mediated passive diffusion 15 . We therefore supposed that there was a limit on the formation of a selection pressure during the process of Acr transmembrane transport in the evolved strain, while the endogenous GSH rapidly detoxified Acr to form the less-toxic Acr-GSH adduct.…”