Ogataea polymorpha
Ogataea angusta ( usually still under the Pichia angusta synonyms or Hansenula polymorpha known) is a methylotrophic yeast species with unusual properties. It is used as a protein factory for pharmaceutical products and is one of a limited number of methylotrophic yeasts (yeasts, which can on methanol, formerly also known as methyl alcohol, grow ).
Basics
Other yeast species with this ability are Candida boidinii, Pichia methanolica and Pichia pastoris. P. angusta belongs taxonomically to the family of Saccharomycetaceae. In recent Taxonomiebüchern the genera Pichia and Hansenula have been combined, and H. polymorpha was renamed in Pichia angusta. According to the desire of many scientists, however, the popular name of H. polymorpha is to be maintained. After but found that Pichia is polyphyletic, the species was placed in the genus Ogataea and is therefore called valid Ogataea angusta. There are three different strains of this type of unexplained relationship and independent origin. They were discovered in soil samples in the gut of insects and rotten orange juice since the beginning of the fifties of the last century. They have different properties and are either popular items of basic scientific research or by genetic modification for the production of proteins ( proteins ) are used:
- Strain CBS4732 ( CCY38 -22 -2; ATCC34438, NRRL Y -5445 )
- Strain DL -1 ( NRRL Y- 7560; ATCC26012 )
- Master NCYC495 (CBS1976; ATAA14754, NRLL -Y 1798).
The strains CBS4732 and NCYY495 can be crossed with each other, strain DL -1 is not interbreed with the other two. The strains CBS4732 and DL -1 are used for the genetic production of proteins, strain NCYC495 is mainly used for studies of nitrate assimilation. The genome of CBS4732 was completely sequenced.
" H. polymorpha "is a thermo- tolerant microorganism - some strains can still grow at temperatures above 50 ° C. In addition, the yeast can assimilate nitrate (this is very unusual for a yeast) and grow on various sugars, glycerol or methanol ( the better-known baker's yeast only grows on glucose and turns it into alcohol). Cells grown at elevated temperature, trehalose accumulate, a sugar that is found in insects, and use it for heat protection. Although trehalose is not necessary for growth, but for the acquisition of thermo - tolerance. The necessary steps for the synthesis of trehalose and yeast species have been elucidated for this TPS1, the gene for the key enzyme in the synthesis method, isolated and characterized. All methylotrophic yeast species have an identical pathway for the use of methanol. The growth on methanol is a massive increase of peroxisomes, certain cell organelles, accompanied ( Fig. 1).
In them the first steps of the methanol metabolism takes place. " H. polymorpha "is a model organism to study the functions of peroxisomes and the functions underlying molecular biology. During growth on methanol certain enzymes of the metabolic pathway are produced in large quantities, such as MOX (methanol oxidase), FMDH ( formate dehydrogenase) and DHAS ( dihydroxyacetone synthase). Their presence is regulated by controlling the transcription (formation of mRNA) of the corresponding genes. In the above-mentioned related yeast species C. boidinii, P. methanolica and P. pastoris, this expression strictly dependent on the presence of methanol, while in " H. polymorpha " this by suitable amounts of glycerol or glucose starvation conditions (glucose starvation ) is caused.
Pichia angusta produced glycoproteins (proteins that are attached to the sugar chain-like molecules), - there are N -and O-linked sugar chains. , The terminal sugar ( mannose ) are linked in chains with N- alpha-1, 2 bonds, and no potentially allergenic alpha-1 ,3- bond, such as in the yeast Saccharomyces cerevisiae.
Pichia angusta is an excellent platform for the production of recombinant proteins, particularly pharmaceuticals such as insulin for diabetics, for hepatitis B vaccines or IFN alpha -2a for the treatment of hepatitis C.
The unusual properties described above make " H. polymorpha " an attractive platform for recombinant production of proteins. Based on genetically modified " H. polymorpha " yeasts there are already numerous products and production processes ( in the title of some important examples are given ). Descendants of the strains CBS4732 and DL -1 are used for this. Other yeast species for this purpose are, inter alia Pichia pastoris, Arxula adeninivorans, Saccharomyces cerevisiae and other (see: protein overexpression in yeast ).
Yeasts are microorganisms that can be used in large fermenters in a short time to cultivate large cell densities. " H. polymorpha " is a safe organism, as it contains no pyrogenic or pathogenic substances. Yeasts could dismiss proteins into the medium ( secrete ), since they have the necessary structures in eukaryotic cells, such as that of humans. The gut bacteria can not do this, for example. " H. polymorpha " provides attractive genetic elements for the efficient production of proteins.
Figure 3 shows the general scheme of a vector is shown ( a circular DNA molecule for transformation of a yeast strain in a genetically modified protein producers). Such a vector (or plasmid designated ) must contain different genetic elements:
Application
The Pichia angusta platform is used by several biotechnology companies to develop production of important proteins, including the company ARTES Biotechnology GmbH in Langenfeld, PharmedArtis in Aachen and the Leibniz Institute of Plant Genetics and Crop Plant Research ( IPK).
HPWN ( Hansenula polymorpha worldwide network)
Many academic research groups worldwide study this organism. In 2000, a scientific society under the name HPWN ( Hansenula polymorpha worldwide network) by Prof. Dr. Marten Veenhuis, Groningen, and Prof Dr Gerd Gellissen, Dusseldorf, was founded. Every two years, organizes scientific meetings.