|Content of Engineering Photosynthetic Pathways
The scientific challenges encountered during the last decade by attempts at
improving photosynthetic productivity, even when successful, generated further
questions, but even the lack of success has taught us many things. As the conclusion
for this chapter, we would like to explore the approaches necessary for future
achievements in improvement of crop productivity.
One most important requisite for manipulating physiology of an organismis to
accumulate information about the precise mechanisms of function of the key
protein(s) or enzyme(s) in question. This includes detailed knowledge on gene
structure and the regulation of gene and protein expression of enzymatic properties
and subcellular location. ADPglucose pyrophosphorylase, for example, had
been studied extensively over a long period, from its biochemistry in vitro through
to regulation of activity in vivo (Preiss et al., 1991). However, only the introduction
of a gene, modified to be insensitive to feedback regulation, into potato tuber
amyloplasts resulted in increased starch synthesis (Preiss, 1996). FBP/SBPase
from a cyanobacterium has been shown to improve productivity in tobacco
(Miyagawa et al., 2001). Since the functional sites of these enzymes are the chloroplast
stroma, the selection of the promoter and the transit sequences for expression of these proteins could easily be accomplished based on previous
knowledge. Another strategy, antisense suppression of resident genes has
revealed the significance of particular enzymes in a postulated metabolic
Similar considerations are also valid for RuBisCO research. We are still
ignorant, for example, about either the residues that determine the Srel
how carbon and oxygen atoms are enabled to overcome spin prohibition on the
RuBisCO protein for the oxygenation of RuBP, and about which residues limit the
reaction rate in overall catalysis (Cleland et al., 1998; Roy and Andrews, 2000).
Translation of rbcL mRNA and association of RuBisCO peptides are important
topics about which not enough is known (Houtz and Portis, 2003; Roy and
Andrews, 2000). In general, the steps of posttranslational folding in plants and
other organisms, whether E. coli, yeast, or human, must become known (Frydman,
2001). RuBisCO should provide an excellent model protein for study, considering
that plants are able to synthesize up to 200 mg/ml of RuBisCO protein within days
during the greening of leaves.
Engineering of the chloroplast genome has become the transformation strategy
that promises to overcome problems encountered in the genetic manipulation
of nuclear chromosomes for functions that must reside in plastids (Daniell, 1999).
The technology will be indispensable for the metabolic engineering of pathways
such as the PCR cycle, and starch and lipid biosyntheses. In this context,
establishing methods for chloroplast genome engineering in the major crop
species is an important priority.
Introduction of the cyanobacterial CO2-pumping system into the plasma membrane
of mesophyll cells or the chloroplast envelope may be one future direction.
Some improvement in the photosynthetic performance of transgenic plants has
already been reported with Arabidopsis (Lieman-Hurwitz et al., 2003).
Interspecies crosses that might lead to the transfer of beneficial genes are not
possible in plants or any higher organism. Attempts at improving physiological
performance in diverse environments can be realized by varying the expression of
genes inherited from the parents. This requires that we understand in more detail
the networks of reactions that constitute the evolutionarily established reaction
bandwidth and allelic plasticity of a species. Science is now beginning to elucidate
the potential of natural intraspecies variation and to probe the upper limits of
plants physiologically, biochemically, and at the molecular genetic levels.
Furthermore, we are learning, as we have pointed out, that it is possible to raise
the potential of organisms and to exceed the intrinsic limits of plant productivity
by introducing genes across species barriers that of a species that cannot be
crossed by traditional breeding.