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Production of heterologous proteins in
Saccharomyces cerevisiae

The availability of efficient vectors has enabled S.
cerevisiaeto be used as a “factory” for the products of
many foreign (heterologous) genes. One of the most
significant products to have emerged from this is a vac-
cine against hepatitis B. It consists of one of the viral
coat proteins, Hepatitis B surface Antigen (HBsAg),
and was produced by transforming yeast cells with
the viral gene encoding this protein. Hepatitis B
vaccine was the first genetically engineered vaccine to
be approved for use in humans (it contains no genes).
Professor Sir Ken Murray was knighted for his work in
developing this vaccine.
Many other proteins have been produced experi-
mentally from yeast, including cellulases, amylases, inter-
feron, epidermal growth factor, and β-endomorphin.
However, there have also been problems in using
Saccharomycesfor heterologous protein production. In
particular, yeast has a relatively poor ability to remove
introns from foreign genes (its own introns are few and
small), so it is most efficient when transformed with
complementary DNA (cDNA) derived in vitrofrom the
messenger RNA of a protein (the introns are spliced out
during the processing of mRNA). Yeast also fails to
recognize the promotor regions of the genes of other
fungi, and it does not always faithfully glycosylate for-
eign proteins. This can be important because several
bioactive proteins are glycoproteins that depend on the
sugar chains for their activity.
These difficulties have served to demonstrate that
S. cerevisiaeis genetically quite different from the
mycelial fungi. For example, E. nidulanscan recognize

the promoter sequences of the genes of other fungi

and also can excise their introns. It may be possible to

use E. nidulansor the fission yeast Schizosaccharomyces

as an alternative to Saccharomycesfor heterologous

protein production. But, in general, it is now thought

that the best approach is to use cell lines related to the

natural producer organism – mammalian cell lines for

mammalian gene products, and so on.

Identification of genes for plant

pathogenicity and differentiation

We saw in Chapter 4 how differentiation-specific

mRNAs were identified in Schizophyllum communeby

comparing the mRNA profiles of cultures grown in con-

ditions where fruitbodies were or were not produced.

The specific mRNAs can then be used as templates to

produce cDNA in vitro. This cDNA, produced from

labeled nucleotides, becomes a probe for binding to

complementary sequences of extracted chromosomal

DNA. In this way a specific gene can be identified

even if nothing is known about the basic genetics of

a fungus. By using such techniques, Wessels and his

co-workers were able to characterize a unique group of

proteins, the hydrophobins, which have major roles in

fungal biology and differentiation. These techniques of

“reverse genetics” have also made it possible to perform

targeted gene disruption. For the fungal gene of

interest, a cDNA is produced in vitroand disrupted to

make it nonfunctional. Then it is transformed into the

fungus, to replace the original gene by homologous

recombination. The following example shows the

power of this technique.

FUNGAL GENETICS, MOLECULAR GENETICS, AND GENOMICS 171

Untranscribed

spacers

(a)

(b)

(c)

(d)

ETS

Pre-rRNA

Mature rRNA

18S 5.8S 28S

ITS

ITS ITS

Processing

ITS

++

18S rRNA 5.8S rRNA 28S rRNA

Fig. 9.10(a–d) Organization and processing
of the eukaryotic rRNA genes.