181 research outputs found
Rubisco function, evolution, and engineering
Carbon fixation is the process by which CO2 is converted from a gas into
biomass. The Calvin Benson Bassham (CBB) cycle is the dominant carbon fixation
pathway on earth, driving >99.5% of the ~120 billion tons of carbon that are
"fixed" as sugar, by plants, algae and cyanobacteria. The carboxylase enzyme in
the CBB, ribulose-1,5-bisphosphate carboxylase/oxygenase (rubisco), fixes one
CO2 molecule per turn of the cycle. Despite being critical to the assimilation
of carbon, rubisco's kinetic rate is not very fast and it is a bottleneck in
flux through the pathway. This presents a paradox - why hasn't rubisco evolved
to be a better catalyst? Many hypothesize that the catalytic mechanism of
rubisco is subject to one or more trade-offs, and that rubisco variants have
been optimized for their native physiological environment. Here we review the
evolution and biochemistry of rubisco through the lens of structure and
mechanism in order to understand what trade-offs limit its improvement. We also
review the many attempts to improve rubisco itself and, thereby, promote plant
growth
Identification of transcriptional activation and inhibitory domains in serum response factor (SRF) by using GAL4-SRF constructs.
The binding of serum response factor (SRF) to the c-fos serum response element has been shown to be essential for serum and growth factor activation of c-Fos. Since SRF is ubiquitously expressed, it has been difficult to measure the activity of SRF introduced into cells. To assay for functions of SRF in cells, we have changed its DNA binding specificity by fusing it to the DNA binding domain of GAL4. Transfection of GAL4-SRF constructs into cells has allowed us to identify SRF's transcriptional activation domain as well as domains which inhibit this activity. First, we found that the transcriptional activation domain maps to between amino acids 339 and 508 in HeLa cells and to between amino acids 414 and 508 in NIH 3T3 cells. Second, we show that in the context of GAL4-SRF constructs, there are two separate domains of SRF that can inhibit its activation domain. Although these domains overlap the DNA binding and dimerization domains of SRF, these functions were not required for inhibition. Finally, we show that one of the inhibitory domains is modular in that it can also inhibit activation when it is moved amino terminal to GAL4's DNA binding domain in an SRF-GAL4-SRF construct. The implications of these inhibitory domains for SRF regulation are discussed
Mutation of serum response factor phosphorylation sites and the mechanism by which its DNA-binding activity is increased by casein kinase II.
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