Romanian Biotechnological Letters Vol. 15, No.

4, 2010
Copyright © 2010 University of Bucharest Printed in Romania. All rights reserved
REVIEW

Methylotrophic yeasts: diversity and methanol metabolism

Received for publication, September 15, 2009
Accepted, Juy 1, 2010

OANA NEGRUŢĂ1*, ORTANSA CSUTAK2, ILEANA STOICA2, ELENA RUSU3,

TATIANA VASSU2
1
University of Bucharest, Faculty of Biology, Center for Research, Consulting and Training in
Microbiology, Genetics and Biotechnology, Aleea Portocalilor 1-3, 060101 Bucharest, Romania
2
University of Bucharest, Faculty of Biology, Department of Genetics, Aleea Portocalilor 1-3,
060101 Bucharest, Romania
3
Titu Maiorescu University, Faculty of Dental Medicine, Street Gheorghe Petrascu, no. 67A,
sector 3, 060101 Bucharest, Romania
*
negruta_oana@yahoo.com

Abstract
Yeast identification has a real importance for selection of technological strains and also for
studying biodiversity and population dynamics in the fermentation systems. Important studies on
yeast population dynamics showed that Saccharomyces genus is dominant in alcoholic fermentation,
while other genera like: Kloeckera, Candida, Pichia, Hansenula, Hanseniaspora and Metschnikowia
are growing in the first step of the process. The purpose of this review was to emphasize the
importance of the methylotrophic yeasts diversity, with the most important morpho-physiological
characteristics, the complexity of the life cycle, and also the main pathway of the methanol
metabolism, with the enzymes involved in this process.

Keywords: methylotrophic yeasts, methanol metabolism, Hansenula polymorpha

Introduction
The yeasts pose great benefits for humankind by their use in the production of food,
wines, beer, and a diversity of bio-chemicals. Preparation of beer and bread with the help of
yeasts has been current practice for millennia. Speaking of food, they can also be used as an
alimentary supplement because of their high content in vitamin B, amino acids and minerals.
At present, they are used not only in the ‘classical’ industrial processes, but also as models for
the study of gene regulation in eukaryotic cells, and also as bio-factories for homologous and
heterologous proteins [7].
Methylotrophic yeasts (belonging to Hansenula, Candida, Pichia, Torulopsis genera)
are capable to metabolise monocarbonic compounds like methanol and formaldehyde [17].
They are very prevalent in nature in particular in moulds, fruit and other vegetable products,
exudates of trees and their barks [6]. A possible explanation for the existence of methylotrophic
yeasts in this kind of habitats is that methanol can derive from the metoxi chain present in lignin
from woods. Almost all tested methylotrophic yeasts are capable to grow on pectic medium, a
polymer rich in the metoxi chain, found especially in fruit [1]. Factors like geographic
localization and the climatic conditions may influence the diversity of yeast [31].
H. polymorpha and P. pastoris are intensively used in the study of biogenesis,
assembly and degradation of the peroxisomes [26]. They were isolated for the first time
decades ago by cultivation on media enriched with methanol [21]. The enzymology of the
methanol dissimilation pathway was elucidated after 10 years and a lot of confusions were
made on the way. The existence of the peroxisomes –induced by methanol in the medium –
which contain the enzymes of methanol metabolism, like: alcohol oxidase (AOX),

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 OANA NEGRUŢĂ, ORTANSA CSUTAK, ILEANA STOICA, ELENA RUSU, TATIANA VASSU

dihydroxyacetone sintase (DHAS) and catalase (CAT) is a very important characteristic of

these yeasts.
H. polymorpha and P. pastoris were successfully used for the production of
heterologous proteins. Moreover, H. polymorpha is the species chosen for studying the
genetic control of enzymes involved in methanol metabolism, nitrate assimilation, resistance
to heavy metals, and to oxidative stress. Also, it has been considered the usage of
methylotrophic yeasts in the treatment of the methanol and formaldehyde containing
wastewater [16, 17].

Alcohol oxidase (AOX)

The first enzyme involved in yeast methanol metabolism, is alcohol oxidase (AOX),
localized in peroxisomes, a particular location [23]. It oxidizes methanol to formaldehyde and
hydrogen peroxide.

Figure. 1. Methanol metabolism pathway in methylotrophic yeasts.

1 – alcohol oxidase, 2 – catalase, 3 – dihydroxyacetone synthase, 4 – formaldehyde dehydrogenase, 5 –
formate dehydrogenase, 6 – dihydroxyacetone kinase, GSH – glutathione, Xu5P – xylulose-5-
phosphate, FBP – fructose-1,6- bisphosphate. [10].

Being one of the key enzymes in methanol utilization, AOX is found in all
methylotrophic yeasts [14]. In its mature active form AOX is a 600kD molecule, consisting of
eight identical subunits of 74kD, with a cvasicubic orientation, each unit including one flavin
adenin dinucleotid molecule (FAD) as prostetic group. The monomers are synthesized in
cytosol and then are post-transcriptional imported into peroxisomes, where their assembly
into the active octamer takes place [11], in contrast with other proteins that are imported as
oligomeres. This might be related to certain favourable conditions met by the peroxisomes
(such as pH value or certain ions concentrations) [30].
AOX is synthesized in high amounts to compensate the low affinity for oxygen, the
AOX promotor being very attractive for the expression of the heterologous proteins, as it is
tightly regulated and very easy induced by methanol [32].
The AOX promoter is one of the most powerful and the most precisely regulated
promotors known [4]. The AOX genes from H. polymorpha, C. boidinii and P. pastoris were
isolated and sequenced. H. polymorpha and C. boidinii possess only one AOX gene while P.
pastoris has two AOX genes (AOX 1 and AOX 2) [23].The AOX gene from H. polymorpha
contains a large promotor region of 1.5kb.
AOX oxidizes not only methanol, but also other alcohols with short chain and
formaldehyde. Therefore, it is used for determination of lower alcohols and formaldehyde, as
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 Methylotrophic yeasts: diversity and methanol metabolism

well as for the construction of sensors that detect alcohol, in conjugation with oxygen and
hydrogen peroxide sensors. It is also very attractive for various biotechnological applications,
including formaldehyde assay in food production, in waste waters and in pharmaceuticals.
The rate of enzymatic oxidation of formaldehyde by AOX from H. polymorpha is twice lower
compared to that corresponding to oxidation of the most appropriate substrate, methanol [27].
In cells, AOX is localized together with dihydroxiacetone synthase and catalase. AOX
and dihydroxiacetone synthase are synthesized in very large amounts, depending on the
growth conditions they can reach a volume of up to 70% of all cellular proteins. Therefore
AOX forms crystalloids which give the characteristic shape of the peroxizomes. Catalase is
located predominately at the periphery of the organelles, between the AOX crystalloids and
the peroxisomal membrane. [34].
Catalase promotes the decomposition of hydrogen peroxide that is a powerful oxidant
agent, toxic for the cells. Catalase is a homotetramer of approximately 241 kDa, which
contains the heme group as cofactor [34]. The absence of catalase from peroxizomes
determines the accumulation of toxic H2O2 and/or other reactive oxygen species (ROS), and
the physiological importance of the enzyme is revealed by the fact that its deficiencies lead to
acatalasemia (a disease which is genetically determined) [13].

The use of formaldehyde by the cells

In methylotrophic yeast metabolism formaldehyde is situated at the branching point of

the assimilation and dissimilation pathways. Because it is a very toxic compound its
intracelular levels are strictly regulated [26].
AOX oxidates methanol to formaldehyde, that is subsequently subjected either to the
action of a transketolase (peroxisomal dihydroxyacetone synthase) or to direct oxidation into
the cytosol. Dihydroxiacetone synthase is a homodimer of 155kDa that catalyses the transfer
of glycoaldehyde from xylulose - 5 phosphate as a donor to the formaldehyde molecule as
acceptor, by a ‘ping-pong’ mechanism. If enough xylulose- 5- phosphate is present into
peroxisomes then formaldehyde is immediately fixed by DHAS, otherwise formaldehyde
diffuses into the cytosol where it is oxidized to CO2 by S- formyl gluthatione hydrolase and
formate dehydrogenase (FDH) [34].
FDH is not essential, but it is still necessary for optimal growth on methanol since it
plays a significant role in formaldehyde detoxification, energy production and the regulation
of the glutathione level in cells [36]. FDH from bacteria and yeasts are some of the most
stable enzymes. They are produced in small amounts. The FDH content from different
methylotrophic bacteria and yeast strains can reach 10 to 18% of the total cellular proteins.
The enzyme is a homodimer of 42kDa with both subunits containing an independent active
centre [7, 19]. Formaldehyde might be oxidized to formate in a glutathione-dependent or
glutathione-independent manner [25]. The enzyme has isoforms, some being NAD+
dependent [33]. FDH hydrolysis S- formylglutathione, resulting the complex enzyme- format,
with the format molecule being subsequently oxidized to CO2. NADH is generated during
formaldehyde oxidation [15].
Pichia angusta
Current name: Pichia angusta (Teun., H.H. Hall & Wick.) Kurtzman, Antonie van
Leeuwenhoek 50(3): 212 (1984)
Pichia angusta has different synonyms: Hansenula angusta Teun., H.H. Hall & Wick.,
Mycologia 52(2): 185 (1961) [1960]; Hansenula polymorpha Morais & M.H. Maia, An. Esc.
Sup. Quim. Univ. Recife 1: 16 (1959) Ogataea polymorpha (Morais & M.H. Maia) Y.
Yamada, K. Maeda & Mikata, Biosc., Biotechn., Biochem. 58(7): 1254 (1994) [37].

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In spite of many descriptions of tested habitats (moulds, rotting fruit and vegetables,
intestinal tract of pigs, soil, residues from the presses of olive oil, milk from cows ill of
mastitis, soil irrigated with treated or untreated wastewater, corn flour, ropy orange juice a.o.)
the complete ecology of this microorganism is still unknown. H. polymorpha presents a
significant capacity to grow at high temperatures. H. polymorpha and C. boidinii as well as
Torulopsis sonorensis (renamed C. sonorensis) were among the first identified, while P.
pastoris was isolated from exudates of trees in North America and Europe.
H. polymorpha is now designated as Pichia angusta with the „polymorpha” epithet
being prioritary - even though it was first used for a species presently known as Ogataea
polymorpha (Moraris and Maia)[20]. The Pichia genus (ex-Hansenula) belongs to
Saccharomycetaceae Family, Hemiascomycetes Order, Ascomycota Fillum. The Pichia genus
species are heterogeneous in some respects, with hat-shaped or spherical ascospores and
different types of Q coenzyme. Pichia genus comprises 92 species being one of the richest
taxons. They can be identified besides classical tests by using RFLP for ITS1, 5.8S, and ITS2 [3].
It has round small cells that may form lateral buds. Ascospores are small, hemispheric
or hat-shaped and connected with each other, or of a planet Saturn form and usually unbound
by fissure of the asca. H. polymorpha can ferment glucose, but not galactose, sucrose, maltose
or raffinose, it does not assimilate carbon under aerobic conditions, and does not form petite
colonies [12]. The life cycle of H. polymorpha is similar to that of S. cerevisiae. It encompasses
a cell proliferation process whereby a cell generates two cells by mitotic division and also
sporulation and mating of the haploids of opposite types (a and α, respectively) [35]. On a rich
medium the haploid and diploid cells germinate. On a minimal (sporulation) medium the
diploid cells form some ascus with four ascospores in meiosis and soprulation.
Pichia pastoris
Current name: Pichia pastoris (Guillierm.) Phaff [as 'pastori'], Antonie van
Leeuwenhoek 22: 115 (1956)
Pichia pastoris has different synonyms: Endomyces pastoris (Guillierm.) Zender [as
'pastori'], (1926), Komagataella pastoris (Guillierm.) Y. Yamada, M. Matsuda, K. Maeda &
Mikata, Biosc., Biotechn., Biochem. 59(3): 444 (1995), Petasospora pastoris (Guillierm.)
Boidin & Abadie [as 'pastori'], Bull. trimest. Soc. mycol. Fr. 70: 365 (1955) [1954],
Saccharomyces pastoris (Guillierm.), Lodder & Kreger-van Rij [as 'pastori'], Yeasts, a
taxonomic study, [Edn 1] (Amsterdam): 197 (1952), Zygosaccharomyces pastoris Guillierm.
[as 'pastori'], C. r. hebd. Séanc. Mém. Soc. Biol. 71: 466-470 (1919), Zygowillia pastoris
(Guillierm.) Kudryavtsev, (1954), Zymopichia pastoris (Guillierm.) E.K. Novák & Zsolt, Acta
bot. hung. 7: 121 (1961) [38].
It has oval small cells with unipolar budding. On solid medium it forms white or
cream-colored, non-filamentous colonies. It grows on various media, on a variety of simple
carbon sources including glucose, glycerol, galactose, fructose, ethanol, methanol, alanine,
lysine, succinate, etyliamine, mannitol, L-rhamnose and trehalose. P. pastoris does not grow
on galactose, arabinose, ribose, maltose, sucrose, raffinose, melibiose, cellulose or starch. P.
pastoris is homotalic, isolation of mutants being very difficult because of this characteristic. It
often exhibits aberrant segregation, and low viability of the spores [22]. This suggests
instability of the mating type which can be used in the process of selection of mutants, as it
offers a chance of detecting some heterotalic strains. [29].
Pichia guiliermondii
Current name: Pichia guilliermondii Wick., J. Bact. 92: 1269 (1966), and synonyms
Yamadazyma guilliermondii (Wick.) Billon-Grand, Mycotaxon 35(2): 203 (1989) [39].
C. guilliermondii is a common species both in natural and clinical environment. Since
the first description in 1917 made by Castellani (Endomyces guilliermondii), a big number of
5372 Romanian Biotechnological Letters, Vol. 15, No. 3, 2010
 Methylotrophic yeasts: diversity and methanol metabolism

new species and varieties phenotipically similar to C. guilliermondii was described. P.

guilliermondii Wickerham represents a collection of sporogenous strains some of them
belonging to the Candida guilliermondii sporogenous species. Therefore each C.
guilliermonidii strain is theoretically capable to hybridise with any strain of P. guilliermondii.
Sources of isolation are very diverse - plants, lake water, cow rumen, soil polluted
with oil [28], it was isolated also from insects on Ulmus sp., unpolluted soil and water [41], and
from shrimp [18] and other invertebrates but its most common habitat is the nectar of flowers
were they are to be found together with Metschnikovia reukaufii. Strains from the sea were
found in low salinity waters (e.g. the Adriatic) but also in sea with hipersalinity together with
strains of Candida, Debaryomyces, and Metschnikowia. It is isolated from marine medium
especially in the rainy seasons [9]. The species has a large geographic distribution [2].
The cells are heterogenous, many of them having an oblong shape (app. 2x10µ),
sometimes forming a pseudomycelium, but not true hyphae. The ascae contain 1-2 hat-shaped
ascospores which enhance their volume after being freed. All natural strains that were
examined or collection strains of P. guilliermondii are heterothalic, mating types being
designed mat+ and mat-. Homothalic strains are not described, but some of the meiotic
segregates of the hybrids obtained by protoplast fusion develop homothalic phenotypes. Such
phenotypes were designated as pseudohomothallic (mat+,-). It was hypothesized that
pseudohomothallic (mat+,-) strains are aneuploids.
Candida boidinii
Current name: Candida boidinii C. Ramírez [as 'boidini'], Microbiol. esp. 6(3): 251
(1953), [40] also can be found under diverse synonyms: Candida koshuensis Yokotsuka & S.
Goto, Candida methanolica Oki & Kounu, Candida olivarium Santa María, Candida
ooitensis Kumamoto & Seriu, Candida queretana T. Herrera & Ulloa, Candida silvicola
Shifrine & Phaff var. melibiosica Nowakowska-Waszczuk & Pietka, Hansenula alcoolica
Urakami, Kloeckera boidinii nom. nud., Torulopsis enokii Urakami (http://www.cbs.knaw.nl).
Yeast strains described as C. boidinii are usually anamorphic. Molecular techniques allow
now the identification of teleomorphic Candida species [8].
It seems that the presence of Candida genus is a very important coastal pollution
marker for marine environments. The isolation and identification of the yeasts from sand and
sea water from Brazil showed the presence of 292 yeast strains belonging to 4 genera and 31
species, with Candida being dominant and many strains of C. boidinii being present [18].
Many strains of C. boidinii were isolated from fermented olives, together with other
representatives of the genus Candida, as well as of Hansenula, Pichia, Torulopsis and
Saccharomyces genera [5].
It has oval small cells with multilateral budding, simple or elaborated pseudohyphae,
white or cream colonies. On solid medium it forms white or cream colonies. It grows on
various media, on a variety of simple carbon sources including galactose, ribose, arabinose,
sucrose, maltose, lactose, L-rhamnose, ethanol, methanol.

Conclusions
In conclusion, the methylotrophic yeasts, and especially H. polymorpha and P.
pastoris are used in the research area and also in biotechnological applications, one of the
most important being the production of heterologous proteins of a large industrial and
medicine importance. The increase of SCP (Single Cell Protein) requirements or the
remediation of the polluted systems by making use of natural alternatives, represent
important reasons for the necessity of characterizing the methylotrophic yeasts.

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References
1. Blanco Pilar, Sieiro Carmen, Villa Tomaès G., - Production of pectic enzymes in yeasts, FEMS
Microbiology Letters, 175, pp: 1-9, 1999
2. Butinar L., Santos S., Martins-Spencer I., Oren A., Cimerman-Gunde N.,– Yeast diversity in
hypersaline habitats, FEMS Microbiology Letters 244, pp: 229–234, 2005
3. Carvajal-Villa Mercedes, Querol Amparo, Belloch Carmela,– Identification of species in the genus
Pichia by restriction of the internal transcribed spacers (ITS1 and ITS2) and the 5.8S ribosomal DNA
gene, Antonie van Leeuwenhoek, 90, pp: 171–181, 2006
4. Cereghino Geoffrey, P. Lin and Cregg James M., - Applications of yeast in biotechnology: protein
production and genetic analysis, Current Opinion in Biotechnology , 10, pp: 422–427, 1999
5. Coton Emmanuel, Coton Monika, Levert Delphine, Casaregola Serge, Sohier Daniele, – Yeast ecology
in French cider and black olive natural fermentations, International Journal of Food Microbiology 108,
pp: 130 – 135, 2006
6. Craveri R., Cavazzoni V., Sarra P. G., Succi G., Molteni L., Cardini G. and D’Fiore,. - Taxonomical
examination and characterization of a methanol-utilizing yeast, Antonie van Leeuwenhoek 42, pp: 533-
540, 1976
7. Cremata José A, – Conventional and non-conventional yeasts in modern biotechnology, Biotecnologia
Aplicada; Vol. 16, No. 2, pp: 117-125, 1999
8. Daniel Heide-M., Sorrell Tania C. and Meyer Wieland, - Partial sequence analysis of the actin gene and
its potential for studying the phylogeny of Candida species and their teleomorphs, International Journal
of Systematic and Evolutionary Microbiology, 51, pp: 1593–1606, 2001
9. De Almeida Joao M.G.C.F.,– “Yeast community survey in the Tagus estuary”, FEMS Microbiology
Ecology 53, pp: 295–303, 2005
10. Gellissen Gerd, Kunze Gotthard, Gaillardin, Claude Cregg, James M., Berardi Enrico, Veenhuis
Marten, Van der Klei Ida, - New yeast expression platforms based on methylotrophic Hansenula
polymorpha and Pichia pastoris and on dimorphic Arxula adeninivorans and Yarrowia lipolytica – A
comparison, FEMS Yeast Research, 5, pp: 1079–1096, 2005
11. Gunkel Katja, Van Dijk Ralf, Veenhuis Marten, and. Van der Klei Ida J, – Routing of Hansenula
polymorpha Alcohol Oxidase: An Alternative Peroxisomal Protein-sorting Machinery, Molecular
Biology of the Cell, Vol. 15, pp: 1347–1355, 2004
12. Hansen Hans and Hollenberg Cornelis P.,–Non-conventional yeasts in Biotechnology. A Handbook, ed.
Springer, pp: 293-298, 1996
13. Horiguchi Hirofumi, Yurimoto Hiroya, Kato Nobuo, and Sakai, Yasuyoshi, - Antioxidant System
within Yeast Peroxisome ; Biochemical and Physiological Characterization of CbPmp20 in the
methylotrophic yeast Candida boidinii”, The Journal of Biological Chemistry Vol. 276, No. 17, pp:
14279–14288, 2001
14. Ito Takashi, Fujimura Shuki, Uchino Masataka, Tanaka Naoto, Matsufuji Yoshimi, Miyaji Tatsuro,
Takano Katsumi, Nakagawa Tomoyuki and Tomizuka Noboru, - Distribution, diversity and regulation
of alcohol oxidase isozymes, and phylogenetic relationships of methylotrophic yeasts, Yeast, 24, pp:
523–532, 2007
15. Jones John G. and Edward Bellion, – Methanol oxidation and assimilation in Hansenula polymorpha,
Biochem. J., 280, pp: 475-481, 1991
16. Kaszycki Paweł, Koloczek Henryk, - Biodegradation of formaldehyde and its derivatives in industrial
wastewater with methylotrophic yeast Hansenula polymorpha and with the yeast-bioaugmented
activated sludge, Biodegradation 13, pp: 91–99, 2002
17. Kaszycki Pawel, Tyszka Małgorzata Malec, Przemysław, Kołoczek Henryk, - Formaldehyde and
methanol biodegradation with the methylotrophic yeast Hansenula polymorpha. An application to real
wastewater treatment Biodegradation 12, pp: 169–177, 2001
18. Kutty Sreedevi N., and Philip Rosamma, – Marine yeasts — a review, Yeast 25, pp: 465–483, 2008
19. Labrou Nikolaos E. and Daniel J. Rigden,– Active-site characterization of Candida boidinii formate
dehydrogenase, Biochem. J., 354, pp: 455- 463, 2001.
20. Meddelhoven Wouter J., – “Hansenula polymorpha Biology and Applications”, Editura Wiley-VCH,
pp: 1 – 5, 2002,
http://books.google.com/books?id=rUc1Cpac8R0C&pg=PP1&dq=hansenula+polymorpha&lr=&hl=ro
&sig=ACfU3U1anXnCmmSqlRWfXQNXw2lQa4apgpawel, accessed at august 2008
21. Murrell J. Colin, Dalton Howard,– Methane and Methanol Utilizers, Ed. Springer, pp: 207-209, 1992.
http://books.google.com/books?id=RXTkMd5W-

5374 Romanian Biotechnological Letters, Vol. 15, No. 3, 2010

 Methylotrophic yeasts: diversity and methanol metabolism

W0C&printsec=frontcover&dq=methane+and+methanol+utilizers&lr=&hl=ro&sig=ACfU3U2PBx5b_
azvsQBE1gffV2DURlpYPg accessed in august 2008
22. Ogrydziak David M., - Development of genetic maps of non-conventional yeasts, J. Basic Microbiol.,
28, pp: 186-196, 1988
23. Ozimek Paulina, Veenhuis Marten and Van der Klei Ida J., - Alcohol oxidase: a complex peroxisomal,
oligomeric flavoprotein, FEMS Yeast Research, 5, pp: 975 – 983, 2005
24. Roa Michele and Blobel Gunter, - Biosynthesis of peroxisomal enzymes in the methylotrophic yeast
Hansenula polymorpha, Proc. Natl. Acad. Sci.USA, vol. 80, pp: 6872- 6876, 1983
25. Sakai Yasuyoshi, Murdanoto Agung Primanto, Konishi Tohru, Iwamatsu Akihiro, and Kato Nobuo, -
Regulation of the Formate Dehydrogenase Gene, FDH1, in the Methylotrophic Yeast Candida boidinii
and Growth Characteristics of an FDH1-Disrupted Strain on Methanol, Methylamine, and Choline,
Journal of Bacteriology, Vol. 179, No. 14, pp: 4480–4485, 1997
26. Sakai Yasuyoshi, Yurimoto Hiroya, Matsuo Hideaki and Kato Nobuo,– Regulation of peroxisomal
proteins and organelle proliferation by multiple carbon Sources in the methylotrophic yeast, Candida
boidinii, Yeast 14, pp: 1175–1187, 1998
27. Shleev S. V., Shumakovich G. P., Nikitina O. V., Morozova, O. V., Pavlishko H. M., Gayda G. Z., and
Gonchar M. V., - Purification and Characterization of Alcohol Oxidase from a Genetically Constructed
Over_producing Strain of the Methylotrophic Yeast Hansenula polymorpha, Biochemistry (Moscow),
Vol. 71, No. 3, pp: 245-250, 2006
28. Sibirny Andrei A., – Non-conventional yeasts in Biotechnology; A Handbook, ed. Springer, pp: 255-
271, 1996
29. Sreekrishana Koti and Kropp Keith E., – Non-conventional yeasts in Biotechnology. A Handbook, , ed.
Springer, pp: 204-250, 1996
30. Stewart Mary Q, Esposito Renee D., Gowani Jehangir and Goodman, Joel M. – “Alcohol oxidase and
dihydroxyacetone synthase, the abundant peroxisomal proteins of methylotrophic yeasts, assemble in
different cellular compartments ”, Journal of Cell Science 114, pp: 2863-2868, 2001
31. Valles Belen Suarez, Pando Bedrinana Rosa, Fernandez Tascon Norman, Querol Simon Amparo,
Rodrıguez, Madrera Roberto, – Yeast species associated with the spontaneous fermentation of cider,
Food microbilogy, 24, pp: 25-31, 2007
32. Szamecz Bela, Urban Gabriella, Rubiera Roger, Kucsera Judit and Dorgai Laszlo, - Identification of
four alcohol oxidases from methylotrophic yeasts, Yeast 22, pp: 669–676, 2005
33. Tishkov V. I. and Popov V. O., – Catalytic Mechanism and Application of Formate Dehydrogenase,
Biochemistry (Moscow), Vol. 69, No. 11, pp: 1252-1267, 2004
34. Van der Klei Ida J., Yurimoto, Hiroya Sakai Yasuyoshi, Veenhuis Marten, - The significance of
peroxisomes in methanol metabolism in methylotrophic yeast , Biochimica et Biophysica Acta 1763,
pp: 1453–1462, 2006
35. Vassu T., Stoica .I, Csutak O., Musat F, - Genetica microorganismelor si inginerie genetica microbiana
- Note de curs si tehnici de laborator, Editura Petriona, pp: 179, 2001.
36. Yurimoto Hiroya, Lee Bumjun, asudaFumi Y, Sakai Yasuyoshi and Kato Nobuo, - Alcohol
dehydrogenases that catalyse methyl formatesynthesis participate in formaldehyde detoxification in the
methylotrophic yeast Candida boidinii, Yeast, 21, pp: 341–350, 2004
37. http://www.indexfungorum.org/Names/SynSpecies.asp?RecordID=107144 accessed at 03.04.2009
38. http://www.indexfungorum.org/Names/SynSpecies.asp?RecordID=303634 accessed at 03.04.2009
39. http://www.indexfungorum.org/Names/SynSpecies.asp?RecordID=337010 accessed at 03.04.2009
40. http://www.indexfungorum.org/Names/SynSpecies.asp?RecordID=344025 accessed at 03.04.2009
41. www.cbs.knaw.nl, accessed at august 2008

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