BLBS102-c33 BLBS102-Simpson March 21, 2012 14:5 Trim: 276mm X 219mm Printer Name: Yet to Come
33 Biochemistry of Beer Fermentation 631of the maltose transporter gene (MALT) with high-copy-number
plasmids in a lager yeast strain has been found to accelerate
the fermentation of maltose during high-gravity (24◦P) brewing
(Kodama et al. 1995). The constitutive expression ofMALSand
MALRhad no effect on maltose fermentability.
The control overMALgene expression is exerted at three
levels. The presence of maltose induces, whereas glucose re-
presses, the transcription ofMALSandMALTgenes (Federoff
et al. 1983a, 1983b, Needleman et al. 1984). The constitutively
expressed regulatory protein (MALR) binds near theMALSand
MALTpromotors and mediates the induction ofMALSand
MALTtranscription (Cohen et al. 1984, Chang et al. 1988,
Ni and Needleman 1990). Experiments withMALR-disrupted
strains led to the conclusion that MalRp is involved in glucose
repression (Goldenthal and Vanoni 1990, Yao et al. 1994). Rela-
tively little attention has been paid to posttranscriptional control,
that is, the control of translational efficiency, or mRNA turnover,
as mechanisms complementing glucose repression (Soler et al.
1987). The addition of glucose to induced cells has been re-
ported to cause a 70% increase in the liability of an mRNA pop-
ulation containing a fragment ofMALS(Federoff et al. 1983a).
The third level of control is posttranslational modification. In
the presence of glucose, maltose permease is either reversibly
converted to a conformational variant with decreased affinity
(Siro and L ̈ovgren 1979, Peinado and Loureiro-Dias 1986) or
irreversibly proteolytically degraded depending on the physio-
logical conditions (Lucero et al. 1993, Riballo et al. 1995). The
latter phenomenon is called catabolite inactivation. Glucose re-
pression is accomplished by the Mig1p repressor protein, which
is encoded by theMIG1gene (Nehlin and Ronne 1990). It has
been shown that Mig1p represses the transcription of all three
MALgenes by binding upstream of them (Hu et al. 1995). The
MIG1gene has been disrupted in a haploid laboratory strain
and in an industrial polyploid strain ofS. cerevisiae(Klein et al.
1996). In theMIG1-disrupted haploid strain, glucose repression
was partly alleviated; that is, maltose metabolism was initiated
at higher glucose concentrations than in the corresponding wild-
type strain. In contrast, the polyploidmig1strain exhibited an
even more stringent glucose control of maltose metabolism than
the corresponding wild-type strain, which could be explained
by a more rigid catabolite inactivation of maltose permease,
affecting the uptake of maltose.
Allα-glucoside transport systems so far characterized in yeast
are H+-transporters that use the electrochemical proton gradient
to actively transport these sugars into the cell (Crumplen et al.
1996, Stambuk and de Araujo 2001). It has been shown that
maltose uptake is the rate-limiting step of fermentation (Kodama
et al. 1995, Wang et al. 2002, Rautio and Londesborough 2003).
At least three different maltose transporters have been identi-
fied inS. cerevisiae, and while theMALx1transporters (and
probably the twoMPH2andMPH3alleles) encode high affinity
(Km=2–4 mmol l−^1 ) maltose permeases, theAGT1permease (a
gene present in partially functionalmal1gloci) transports mal-
tose with lower (Km≈20 mmol l−^1 ) affinity (Han et al. 1995,
Stambuk and de Araujo 2001, Day et al. 2002a, Alves et al.
2007, 2008).AGT1is found in manyS. cerevisiaelaboratory
strains and maps to a naturally occurring, partially functionalallele of theMAL1locus (Han et al. 1995). Agt1p is a highly
hydrophobic, postulated integral membrane protein. It is 57%
identical to Mal61p (the maltose permease encoded atMAL6)
and is also a member of the 12 transmembrane domain super-
family of sugar transporters (Nelissen et al. 1995). Like Mal61p,
Agt1p is a high-affinity, maltose/proton symporter, but Mal61p is
capable of transporting only maltose and turanose, while Agt1p
transports these twoα-glucosides as well as several others in-
cluding isomaltose,α-methylglucoside, maltotriose, palatinose,
trehalose and melezitose.AGT1expression is maltose inducible
and induction is mediated by the Mal-activator.
Brewing strains of yeast are polyploid, aneuploid, or, in the
case of lager strains, alloploid. Recently, Jespersen et al. (1999)
examined 30 brewing strains of yeast (5 ale strains and 25 lager
strains) with the aim of examining the alleles of maltose and
maltotriose transporter genes contained by them. All the strains
of brewer’s yeast examined, except two, were found to contain
MAL11andMAL31sequences, and only one of these strains
lackedMAL41.MAL21was not present in the 5 ale strains
and 12 of the lager strains.MAL61was not found in any of
the yeast chromosomes other than those known to carryMAL
loci. Sequences corresponding to theAGT1gene (transport of
maltose and maltotriose) were detected in all but one of the
yeast strains.
Although maltose is easily fermented by the majority of yeast
strains after glucose exhaustion, maltotriose is not only the least
preferred sugar for uptake by theseSaccharomycescells, but
many yeasts may not use thisα-glucosidase at all (Zheng et al.
1994a, Yoon et al. 2003). Incomplete maltotriose uptake dur-
ing brewing fermentations results in yeast fermentable extract
in beer, material loss, greater potential for microbiological sta-
bility and sometimes atypical beer flavor profiles (Stewart and
Russell 1993). Maltotriose uptake from wort is always slower
with ale strains than with lager strains under similar fermenta-
tion conditions. However, the initial transport rates are similar
to those of maltose in a number of ale and lager strains. El-
evated osmotic pressure inhibits the transport and uptake of
glucose, maltose and maltotriose with maltose and maltotriose
being more sensitive to osmotic pressure than glucose in both
lager and ale strains. Ethanol (5% w/v) stimulated the transport
of maltose and maltotriose, due in all probability to an ethanol-
induced change in the plasma membrane configuration, but had
no effect on glucose transport. Higher ethanol concentrations
inhibited the transport of all three sugars.
Maltotriose uptake shows complex kinetics indicating the
presence of high- and low-affinity transport activities, and stud-
ies on sugar utilization revealed that maltose and maltotriose
are apparently transported by different permeases (Zheng et al.
1994b, Zastrow et al. 2001). Two genes are recognized as per-
mease genes for transporting maltotriose in yeasts:AGT1trans-
porter fromS. cerevisiaeandMTY1(also calledMTT1) perme-
ase fromS. pastorianus. They are characterized as low-affinity
(Km≈20 mmol l−^1 ) maltotriose transporters (Stambuk and de
Araujo 2001, Dietvorst et al. 2005, Salema-Oom et al. 2005,
Alves et al. 2007, 2008).
Recent reports regarding the observed patterns of maltose and
maltotriose utilization by yeast cells show contradicting results.