Applied Usage of Yeast Spores as Chitosan Beads

Abstract

bIn this study, we present a nonhazardous biological method of producing chitosan beads using the budding yeast Saccharomyces cerevisiae. Yeast cells cultured under conditions of nutritional starvation cease vegetative growth and instead form spores. The spore wall has a multilaminar structure with the chitosan layer as the second outermost layer. Thus, removal of the outermost dityrosine layer by disruption of the DIT1 gene, which is required for dityrosine synthesis, leads to exposure of the chitosan layer at the spore surface. In this way, spores can be made to resemble chitosan beads. Chitosan has adsorptive features and can be used to remove heavy metals and negatively charged molecules from solution. Consistent with this practical application, we find that spores are capable of adsorbing heavy metals such as Cu 2؉ , Cr 3؉ , and Cd 2؉ , and removal of the dityrosine layer further improves the adsorption. Removal of the chitosan layer decreases the adsorption, indicating that chitosan works as an adsorbent in the spores. Besides heavy metals, spores can also adsorb a negatively charged cholesterol derivative, taurocholic acid. Furthermore, chitosan is amenable to chemical modifications, and, consistent with this property, dit1⌬ spores can serve as a carrier for immobilization of enzymes. Given that yeast spores are a natural product, our results demonstrate that they, and especially dit1⌬ mutants, can be used as chitosan beads and used for multiple purposes.

Bioconversion of d-glucose to d-psicose with immobilized d-xylose isomerase and d-psicose 3-epimerase on Saccharomyces cerevisiae spores

“…To eliminate the glycosylation effects on the activity of DPEase, two asparagines of potential N-glycosylation sites were substituted with glutamine by site-directed mutagenesis, but mutant DPEase still showed no activity. In our previous study, β-galactosidase was successfully immobilized on the chitosan layer of dit1Δ spores by chemical method [35]. In contrast to chitosan chemically obtained, dit1Δ spores could function as natural chitosan beads, which could be easily obtained with low cost.…”

“…In our previous work, we successfully developed two new methods of enzyme immobilization based on the structure of yeast spore wall [28,35]. For the biological approach, the target enzyme was expressed in sporulating cells and the outmost layer dityrosine can function as a barrier to prevent the diffusion of soluble proteins [29].…”

Emerging Biomedical Applications of Nano-Chitins and Nano-Chitosans Obtained via Advanced Eco-Friendly Technologies from Marine Resources

“…( a ) The study of the enzymatic deacetylation of n-chitin has not been undertaken so far, notwithstanding the fact that engineered deacetylases are available today [ 205 ]. No information exists on the behavior of deacetylases on n-chitin; ( b ) Serratia marcescens B742 mutants were prepared to improve the deproteination of shrimp shell powders, in fact 91.4% was achieved after three days of fermentation [ 206 ]; ( c ) Yeast spores can be deprived of their outermost dityrosine layer by genetic engineering, thus exposing their chitosan layer which becomes available for collection of metals, enzymes, sterols, and for use in medication [ 207 ]; ( d ) Certain agroindustrial discards (corn steep liquor and molasses) can be converted into chitosan by Rhizopus arrhizus and Cunninghamella elegans [ 208 ]; ( e ) The fisheries themselves have much to gain by using n-chitin and n-chitosan for the improved preservation of crustaceans [ 209 ]; ( f ) The treatment of fresh by-products from the canning factories should be revised in the light of existing advanced technologies [ 210 ].…”

β‐1,6‐glucan synthesis‐associated genes are required for proper spore wall formation in Saccharomyces cerevisiae

“…For deletion of BIG1 , KRE6 and KRE1 , the primer pairs HP5 and HP6, HP19 and HP20, HP27 and HP28, were used to generate knockout cassettes, respectively, and pFA6a‐KanMX6 was used as a template. big1∆dit1∆ double mutant was constructed based on the big1∆ haploid cells; deletion of DIT1 was performed as reported before (Zhang et al ., ). To construct big1∆chs3∆ double mutant, haploid big1∆ and chs3∆ (Coluccio et al ., ) cells were crossed.…”

“…Yeast cells were sporulated as described before (Zhang et al ., ). Briefly, yeast cells derived from a single colony were cultured overnight in 5 mL of SD liquid medium with appropriate supplemental amino acids.…”