Pichia stipitis

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Classification

Figure 1. Pichia Stipitis. "Pichia stipitis morphology under various conditions. (A) Pichia stipitis growing exponentially with bud scars; (B) P. stipitis hat-shaped spores seen from top and side; (C) Pseudomycelia formed under carbon-limited continuous culture. (Credit: Photo by Thomas Kuster, USDA, Forest Service, Forest Products Laboratory)" Image and caption found at Sciencedaily.com [1]

Eukarya; Fungi/Metazoa; Fungi; Dikarya; Ascomycota; Saccharomycotina; Saccharomycetes; Saccharomycetales; Debaryomycetaceae; Scheffersomyces

Species

NCBI: Scheffersomyces Stipitis Taxonomy

Scheffersomyces Stipitis

Description and Significance

The fungus P. stipitis is a yeast from the xylose fermenting clade of the Schefferomyces genus. As with most members of the Saccharomycetales order a single P. stipitis individual has a size of 3 to 5 µm in diameter. Budding from asexual reproduction may cause deviation from the usual spherical or ellipsoidal shape of an individual. Under study it has been noted P. stipitis exhibits a cream-colored, smooth-shaped colony [5]. A suitable habitat for P. stipitis includes damp or wet areas rich in organic biomass [8]. Hardwood forests or areas high in agricultural waste are common environments. Temperatures at which growth occurs are 25°C to 37°C with budding most commonly occurring at 25°C. Studies have shown growth ceases at temperatures around 40°C to 45°C [5].

The significance of P. stipitis to industrial-related purposes comes from the ability to ferment the sugar, xylose [4]. Degrading biomass to ethanol at high yields in respect to other ethanol-producing microorganisms has made P. stipitis an important constituent of the alternative fuel and bioenergy industries [7].

Genome Structure

The genome P. stipits is composed of eight linear chromosomes. The total genome is comprised of 15.4 Mbp with chromosomes ranging in size from 3.5 to 0.97 Mbp [9]. Studies have shown that P. stipitis is haploid, because when compared to similar yeasts, who have determined to be haploid, they share a high frequency of recessive mutations [11].

Analysis of the genome sequence have revealed many sequences that code for putative xylose transporters similar to those found in related yeasts, D. hanseii and C. intermedia. In addition to containing genes required for the glycolysis, the tricarboxylic cycle, and the oxidative pentose phosphate pathway, it contains genes for xylose assimilation and ethanol production [9]. Genome sequencing and analysis have led to many possible insights into the metabolism of P. stipits as indicated by the following assertion from T. W. Jeffries et al., "The presence of numerous genes for endoglucanases and Beta-glucosidases, along with xylanase, mannanase, and chitinase activities indicate that it could metabolize polysaccharides in the beetle gut" [9]. The ability of P. stipitis metabolize the polysaccharides found in the beetle gut is not that surprising since the strain was originally isolated from insect larvae belonging to passalid beetles [6]. However, the yeast has the highest known capacity to ferment xylose to ethanol [12], an abundant sugar in the polysaccharides that compose hardwood and other plant material [10]. This aspect of the yeast's metabolism and other reported pathways, such as the fermentation of cellobiose to ethanol [13], have lead to interest in the possibility of using P. stipitis to produce ethanol for use as a biofuel.

Cell Structure, Metabolism and Life Cycle

Pichia stipitis has been shown to be haplotonic and homothallic [11] and goes through ascomycetous sexual reproduction [5].

Pichia stipitis is known for its ability to produce ethanol by fermenting xylose [19]. Xylose enters the cell through a proton symport transporter [16]. A xylose reductase then reduces xylose to xylitol, which is consequently oxidized to xylulose with a xylitol dehydrogenase. Xylulose is phosphorylated and introduced into the pentose phosphate pathway, which will yield three and six carbon sugars that can be utilized in glycolysis [14].

P. stipitis will readily use oxygen as a terminal electron acceptor during catabolysis of sugars, but does not produce ethanol when grown aerobically. When grown under oxygen-limited conditions P. stipitis will begin fermenting xylose and produce ethanol, but this method is inefficient so the cell gains less energy and grows slower [18]. It is possibly for this reason, that the xylose transport system has evolved to be the rate limiting step of xylose fermentation under anaerobic conditions [16]. Oxygen is believed to activate the transport system increasing the rate of xylose up-take [17]. This might be because xylose transport is an energy consuming process so energized oxygenated cells would be able to transport more xylose than less energized anaerobic cells [15], and thus oxygen indirectly influences xylose up-take [17]. This leads to a problem in using P. stipitis for industrial ethanol production since it grows well and transports xylose well under oxygenated conditions, but ferments xylose and produces ethanol well only under anaerobic conditions. This may be overcome through genetic engineering of P. stipitis and its genes.

Ecology and Pathogenesis

Figure 2. Carbon Dioxide Cycle. Biogeochemical significance and environmental contribution: Bioethanol conversion from lignin and cellulose made possible by Pichia Stipitis.. Image from [2]

The fermentation capabilities of P. stipitis are stimulated in environments where oxygen becomes a limitation [7]. Aside from typical forest or agricultural habitats P. stipitis also forms an endosymbiotic relationship with passalid beetles [6]. In the case of a study by Blackwell and Nguyen (unpublished) 400 of the 400 tested passalid beetles were found to contain the microorganism in their guts' [8]. The oxygen-limited digestive tract and hindgut environment of the wood-ingesting beetle permit the degradation of xylose by P. stipitis [4].

It has also been determined P. stipitis can be utilized to ferment glucose from manufactured lumber products. Construction waste in landfills rich in glucose content may contribute to the bioenergy feedstock via the microbes' biomass conversion capabilities. Ethanol yield efficiencies can be as high as 84.7 to 90.7% per unit substrate consumption [6]. Due to the fermentative ability there is potential to utilize food and municipal wastes as well as agricultural and forest residues to significantly contribute to ethanol feedstocks [6].

References

[1] National Center for Biotechnology Information. (2011). Taxonomy Browser. Retrieved Apr 2011, from NBCI: http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?id=4924

[2] Haeng Cho, D. et al. (2010). "Ethanol production from acid hydrolysates based on the construction and demolition wood waste using Pichia stipitis." Bioresource Technology. Volume 102. p. 4439-4443.

[3] NBCI. (2011, Jan). Taxonomy Browser. Retrieved feb 2011, from NBCI: www.fdinodsin.com

[4] Suh, S., et al. (2004). "The beetle gut : a hyperdiverse source of novel yeasts". The British Mycological Society. Volume 109. p. 261-265.

[5] International Mycological Association. (2011). Yeast Species Database. Retrieved Apr 2011, from MycoBank: http://www.mycobank.org/yeast/BioloMICS.aspx?Link=T&DB=0&Table=0&Descr=Pichia%20stipitis&Fields=All&ExactMatch=T

[6] Suh, C., Marshall J., McHugh, J. V., Blackwell, M. (2003). "Wood ingestion by passalid beetles in the presence of xylose-fermenting gut yeasts". Molecular Ecology. Volume 12. p. 3137-3145. http://si-pddr.si.edu/jspui/bitstream/10088/6710/1/Suh_Marshall_McHugh_and_Blackwell.pdf

[7] Joint Genome Institute. (2011). Pichia Stipitis v2.0. Retrieved Apr 2011, from JGI: http://genome.jgi-psf.org/Picst3/Picst3.home.html

[8] Blackwell, Meredith, Cletus P. Kurtzman, Marc-André Lachance, and Sung-Oui Suh. 2009. Saccharomycotina. Saccharomycetales. Version 22 January 2009. http://tolweb.org/Saccharomycetales/29043/2009.01.22 in The Tree of Life Web Project, http://tolweb.org/

[9] Thomas W Jeffries, Igor V Girgoriev, Jane Greenwood, et al. 2007. "Genome sequence of the lignocellulose-bioconverting and xylose-fermenting yeast Pichia stipits". Nature Biotechnology. Volume 25. p. 319-326. http://www.nature.com/nbt/journal/v25/n3/abs/nbt1290.html

[10] Jeffries, T.W. "Engineering yeasts for xylose metabolism". Curr. Opin. Biotechnol. 17, 320-326 (2006). http://www.sciencedirect.com/science?_ob=MImg&_imagekey=B6VRV-4K07FNM-1-7&_cdi=6244&_user=1111158&_pii=S0958166906000668&_origin=search&_coverDate=06%2F30%2F2006&_sk=999829996&view=c&wchp=dGLzVlz-zSkzS&md5=fa9a7ce4d136c69fe930792d7dcc4847&ie=/sdarticle.pdf

[11] Melake, T., Passoth, V. V. and Klinner, U. 1996. "Characterization of the genetic system of the xylose-fermenting yeasts". Antonie Van Leeuwenhoek. Volume 57. p. 215-222.

[12] van Dijken, J. P., van den Bosch, E., Hermans, J. J., de Miranda, L. R. and Scheffers, W. A. 1986. "Alcoholic fermentation by 'non-fermentive' yeasts. Yeast. Volume 2. p. 123-127. http://onlinelibrary.wiley.com/doi/10.1002/yea.320020208/pdf

[13] Parekh, S. R., Parekh, R. S., and Wayman, M. 1988. "Fermentation of xylose and cellobiose by Pichia stipitis and Brettanomyces clausenii". Appl. Biochem. Biotechnol. Volume 18. p. 325-338. http://www.springerlink.com/content/f15751524621628u/

[14] Barnett, J. A. 1976. "Utilization of sugars by yeasts". Adv. Carbohydr. Chem. Biochem. Volume 32. p. 125-234. http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B7CS1-4S934RF-7&_user=1111158&_coverDate=12%2F31%2F1976&_rdoc=1&_fmt=high&_orig=gateway&_origin=gateway&_sort=d&_docanchor=&view=c&_searchStrId=1729124276&_rerunOrigin=google&_acct=C000051676&_version=1&_urlVersion=0&_userid=1111158&md5=ef463a99150ccaa0dfa0b3a7be019868&searchtype=a

[15] Does, A. L. and L. B. Bisson. 1989. "Characterization of xylose uptake in the yeasts Pichia heedii and Pichia stipitis". Appl. Environ. Microbiol. Volume 55. p. 159-164. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC184071/pdf/aem00094-0179.pdf

[16] Kilian, S. G. and N. van Uden. 1988. "Transport of xylose and glucose in the xylose fermenting yeast Pichia stipitis". Appl. Microbiol. Biotechnol. Volume 17. p. 545-548. http://www.springerlink.com/content/pw51576xmp367058/

[17] K. Skoog and B. Hahn-Hagerdal. 1990. "Effect of oxygenation on xylose fermentation by Pichia stipitis". Applied and Environmental Microbiology. Volume 56. p. 3389-3394. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC184958/pdf/aem00092-0167.pdf

[18] Rizzi, M., C. Klein, C. Schultze, N. Bui-Thanh, and H. Dellweg. 1989. "Xylose fermentation by yeasts. 5. Use of ATP balances for modeling oxygen limited growth and fermentation with the yest Pichia stipitis with xylose as a carbon source". Biotechnol. Bioeng. Volume 34. p. 509-514. http://onlinelibrary.wiley.com/doi/10.1002/bit.260340411/abstract

[19] Toivola, A., D. Yarrow, E. van den Bosch, J. P. van Dijken, and W. A. Scheffers. 1984. "Alcoholic fermentation of D-xylose by yeasts". Appl. Environ. Microbiol. Volume 47. p. 1221-1223. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC240200/pdf/aem00163-0035.pdf

Author

Page authored by Josh Wolter and John Wright, students of Prof. Jay Lennon at Michigan State University.

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