Use of yeast spores for microencapsulation of enzymes.
Here, we report a novel method to produce microencapsulated enzymes using Saccharomyces cerevisiae spores. In sporulating cells, soluble secreted proteins are transported to the spore wall. Previous work has shown that the spore wall is capable of retaining soluble proteins because its outer layers work as a diffusion barrier. Accordingly, a red fluorescent protein (RFP) fusion of the α-galactosidase, Mel1, expressed in spores was observed in the spore wall even after spores were subjected to a high-salt wash in the presence of detergent. In vegetative cells, however, the cell wall cannot retain the RFP fusion. Although the spore wall prevents diffusion of proteins, it is likely that smaller molecules, such as sugars, pass through it. In fact, spores can contain much higher α-galactosidase activity to digest melibiose than vegetative cells. When present in the spore wall, the enzyme acquires resistance to environmental stresses including enzymatic digestion and high temperatures. The outer layers of the spore wall are required to retain enzymes but also decrease accessibility of the substrates. However, mutants with mild spore wall defects can retain and stabilize the enzyme while still permitting access to the substrate. In addition to Mel1, we also show that spores can retain the invertase. Interestingly the encapsulated invertase has significantly lower activity toward raffinose than toward sucrose.This suggests that substrate selectivity could be altered by the encapsulation.
Libing
L
Shi
Zijie
Z
Li
Hiroyuki
H
Tachikawa
Xiao-Dong
XD
Gao
Hideki
H
Nakanishi
D001709:Biotechnology; D002473:Cell Wall / Q000201:enzymology / Q000235:genetics; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D013172:Spores, Fungal / Q000201:enzymology* / Q000235:genetics; D000519:alpha-Galactosidase / Q000235:genetics / Q000378:metabolism*; D000097573:Red Fluorescent Protein
D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D000519:alpha-Galactosidase; D000097573:Red Fluorescent Protein