Heat stress on plant reduces photosynthesis and yield. Sharkey 2005 suggested that this effect is due in activation of ribulose bisphosphate
carboxylase oxygenase rubisco, the enzyme responsible for carbon dioxide fixation by moderate heat stress. While Ortiz and Cardemil 2001 found that heat
stress, cause dissociation of LHC and PSII reaction centre. While damage due to ice is well known as irreversible damage of cellular micro architecture that
happen when transition phase from liquid to ice Bryant et al 2001.
2.2 The character of trehalose and its role in several organism
Trehalose α
-D-glucopyranosyl[1-1]- α
-D glucopyranose is a non reducing disaccharide consists of 2 glucose joined with glycosidic bond. This
sugar is ubiquitously found in biological world, such as in bacteria, yeast, fungi, some animals such as insects and worms, and, in lower kingdom of plant e.g. in
fern Selaginella lepidophylla called “resurrection plant” Zentella et al 1999. Recently, however, trehalose is also found in higher kingdom of plants of in a
very small amount. Trehalose is commonly found in spores, fruiting body, seeds, and vegetative tissue. Calaco et al 1995 suggested that this compound is stable
in high temperature and low pH. It is stable and stabilizing sorounding compound against heat Singer and Lindquist 1998; Reinders 1999; Kandror 2003, cold,
drought and salt Garg et al 2002, Jang et al 2003, El-bashiti 2003, oxidative stress Franco et al. 2000; Filinger 2001; Benaorouj et al. 2002 and pressure
Iwahashi et al. 2000. More detail is discuss in the nex subheading. One of the stabilization effects of trehalose on sorounding molecules is
the capability form hydrogen bonding to the molecules. During the absence of water, trehalose is able to form energetically stable conformations bridging with a
number of lipid molecules Chandrasekhar and Gaber, 1988. Calculating the H- bond and the member of molecules of lipid that interact with trehalose at multiple
H-bonding, one molecule trehalose could make 8 hydrogen bonding although some no and less hydrogen bonding were also observed. Furthermore, one
molecule trehalose could interact with 3 molecules of lipid. When the temperature increase lipid molecule is expanded and trehalose intercalates within the
molecule and forms H-bonding. Study at atomic level on trehalose membrane interaction showed that trehalose interact directly to membrane via hydrogen
bonding and at elevated temperature, the stabilization effect is correlated with a
number of trehalose molecules bridging three or more lipid molecules Pireira et al.
2004. This compound has several roles in organisms; however, it may has
different function among various organisms for review, Albein et al. 2003. Trehalose function as energy reservation, e.g on Neurospora tetrasperma which
contains 10 of it’s dry weight on germination while in adult insect, trehalose is also used as energy such as for flying Albein, 2003. On plant material, despite
of osmotic protecting agent, trehalose is also functioning as regulator for glucose, abscisic acid and stress signaling Avonce et al. 2004 that implicated on carbon
allocation and involve in sugar metabolism Vogel et al. 1998; Muller et al. 1999; Garg et al. 2002; Jang et al. 2003. This also has a role in growth and
development of Arabidopsis thaliana Shluepmann et al 2003; 2004. Arabidopsis thaliana
trehalose phosphate synthase AtTPS1 is up regulated along with seed developmental stage and required for full expression of seed maturation marker
gene Eastmond et al. 2002; Schluepmann et al. 2004 and knocked out AtTPS1 is embryo lethal.
Trehalose metabolism also affects biosynthesis and starch degradation. Feeding with trehalose induced starch accumulation and ADP-glucose
pyrophosphorylase gene ApL3 expression Wingler et al. 2000, which lead to
metabolically available carbon immobilization from source to sink that in turn cause growth arrest. The starch induction in chloroplast occurred via
posttranslational redox activation of ADP-glucose pyrophosphorylase by accumulation of trehalose-6-phosphate T6P Kolbe et al. 2005. This growth
arrest is relieved when sugars are added simultaneously together with trehalose Schluepmann et al. 2004. Trend reduction of hexose phosphates activity was
also shown on plant with accumulation of T6P in Arabidopsis Schluepmann et al. 2004. While in yeast, this accumulation causes significant growth reduction
Bonini et al. 2003. Its intermediate product of trehalose in Arabidopsis T6P is needed in small amount for embryo development but in high amount resulting in
growth arrest. Abolish of its synthesis cause embryo lethal Schluepmann et al. 2005.
2.3 Trehalose and its role as stress protecting agent In yeast, trehalose is not synthesized during the exponential phase, but
during stationary phase and sporulation. This sugar is not used until desperate
situation to avoid starvation, and is not used as energy reserve Wiemkem A, 1990. Mutants of E. coli that unable to synthesis trehalose were osmotically
sensitive to glucose medium Giaever et al. 1988. While Alarico et al. 2003 reported that disrupted gene for trehalose synthase in Thermus thermophylus
reduced the capability to survive on salinity condition with maximum concentration of 5. This situation was relieved when trehalose was added into
the medium, while it was not releaved when glycine betaine, manosylglycerate, maltose, or glucose was added to the medium. Trehalose content on stressed
sensitive wheat species has lower level than the resistant one either in normal conditions or in drought and salt El-Bashiti, 2003.
Trehalose is the most effective compatible solute among sugars Sampedro 1988. The effectiveness of trehalose in preserving biomoleculs is related with the
flexibility between the two monomers compared with other sugars such as sucrose and maltose Crowe et al. 1983 by which it conforms to irregular polar
groups of macromolecules provided a better interaction with them. Hence, trehalose could reduce these effects of multiple stresses such as water status
and temperature Kandror et al. 2002, salinity Alarico et al. 2005, oxidative stress Peral et al. 2002, osmotic and oxidative stress Giaever et al, 1988,
Franco et al 2000; Fillinger et al. 2001; Benaorouj 2002 and nutrient starvation. Membrane damage for instance, has been effectively protected from temperature
Macdonald and Johari 2000; Pereira, 2004; Hincha et al 2004; Patist et al. 2005; from freezing and hydration force Yoon et al. 1998 and from water stress
Chen et al. 2001; Bryant et al. 2001 by trehalose. Shinohara et al. 2002 found that trehalose is also involved in cicardian regulation of stress responses and
development. There are three hypothesis was proposed to explain the protective effect of
sugars including trehalose to biological compounds. First, trehalose as water replacement, this theory suggested that trehalose can substitute water molecule
when water is very scare. Trehalose makes hydrogen bonding around the polar and charged group of membrane and protein; hence their native structure is
stabilized Crowe et al. 1997; Carpenter et al. 1994. Second theory, trehalose as water entrapment, suggested that trehalose concentrates water surround the
biological molecules, thereby maintaining their salvation and native structure. Third, sugars as vitrifying agent, by which it making glassy matrix that
mechanically stabilize their native structure Sun et al. 1996. The third
suggestion, however, it is depending on the condition of biological compound when vitrification occur Bryant et al. 2001.
There are many suggestions supporting the theory. Trehalose has a tendency to make a direct interaction with biological compounds, such as
membrane, protein and nucleic acid and prevents their biological damage Luzardo et al. 2000; Ekdawi, 2001; Bordat et al. 2004. This is due to the
capability of trehalose to adsorb water and reduces its dynamic, replaces the hydrogen bond, reduces molecule expansion, reduces over vibration specific site
of molecules. Furthermore, trehalose is very stable of its own molecule and stabilizes its surrounding compounds, as known as biological stabilizer.
Trehalose interacts with phosphate group laid between the head and the tail, keeping the membrane fluidity Crowe et al. 1987; Rudolf et al. 1990. Pirera et
al. 2004 have reported interaction simulation between membrane bilayers
Figure 1. The increase temperature from 325K to 475K increase structural disorder of the membrane. The presence of trehalose 2 mM much reduced the
degree of disorder. Bryant et al. 2001 suggested that the main effect of cytoplasmic small
solutes is simply related with water relation. Solutes increases the osmotic pressures that reduce water removal from the cell and solute also have
volumetric effect that intercalate between lipids, hence, force of hydration is reduced. However, they agree that there is no controversial suggestion about the
first and second theory of biological compounds stabilization by sugars, especially membrane via this manner. For the third theory, however, they suggest
that it is still confusing since the effect of vitrification differs in different situation and different solute used such as the size. The ability of trehalose in stabilizing
the biological compound from freeze is related with the capability of trehalose to disrupt tetrahedral hydrogen bond network of water, and reduce the freezable
water Crowe et al. 1983. Such interesting though is suggested by Alpert 2006, suggesting that stress
tolerant organism is very rare found on multi cellular organism but it does widelyspread on those 5 mm in size. He suggested that there is a trade-off
between desiccation tolerance and growth. Recent molecular and biochemical
research shows that organisms tolerate desiccation through a set of mechanisms,
including sugars that replace water and form glasses that stabilize
F
Figure 1: The damaging effect of heat to membrane and the role of trehalose to decrease the effect. Diagram of membrane bilayer A. Effect of heat to
membrane structure B. Schematic diagram of liquid phase of biological membrane C. Schematic diagram of membrane in gel phase when subjected to
stress D. Schematic diagram of lipid as part of membrane when it subjected to severe stress E. A model of trehalose that is suggested to interact with
membrane at hydrophyllic part of lipid F. This picture is cited from Web www.agronomy.psu.edu, 07252004.
475 K
+ 1mM Trehalose Pereira, 2004
+ 2mM Trehalose
375 K
475 K
A B
C D
E
macromolecules such as proteins and membrane, and via production of anti- oxidants that counter damages resulted from reactive oxygen species. These
protections are often induced by drying, and some of the genes involved may be
homologous in microbes, plants, and animals. As mentioned above that trehalose is also involve in preserving biomolecules
from cold, hence it is well used as cryopreservation agents. Additional of sugars or salts to the solution distort the water structure where which these compounds
may enhance the tetrahedral coordinated hydrogen bond structure or reduced it that respectively called as water structure maker and water structure breaker.
Preservation of biological membrane would be effective with additional of water solute act as water structure breaker that reduced freezable water Branca et al
1999. Including in water structure breaker are ClO
4 -
, MnO
4 -
, Br
-
, Cl
-
, K
+
, Cs
+
, sugars and I
-
. Those belong to water structure maker are Li
+
, Cu
+
, Al
+
, Mg
+
, and OH
-
. While Na
+
, Ag
+
and Ba
+
are in borderline Bryant et al. 2001. Miller and Pablo 2000 suggested heat solution value of trehalose, maltose and sucrose as
water structure breakers. The conformation of sugars in the solution release heat enthalpy, where 19.1, 15.6 and 5.95 kJmol of trehalose, maltose and sucrose
respectively that in turn reduce freezable water. Trehalose is also associated with resistance of organism to oxidative stress,
such to super oxide. Yeast cell that exposed to mild heat stress induced accumulation of trehalose and its resistance to hydrogen peroxide. Conversely,
those lacking of trehalose synthase were sensitive to hydrogen peroxide as its protein oxidation run faster Benarouj et al. 2001. Trehalose is also found to be
essential for mycolic acid biosynthesis in Corrynebacterium glutamicum. The absence of mycolic acid in defective trehalose synthase mutant caused cell wall
disorder, excretion of amino acids and impairment with bacterial growth Wolf 2002.
There is no discussion about trehalose in combating the deleterious effect of high salinity has been reported. However, Garg et al. 2002 and Jang et al.
2003 found that transgenic rice bearing gene for trehalose synthase fusion of TPS1-TPP
were also tolerant to salt and cold, despite to drought.
2.4 Other compatible solutes