In yeast, galactose triggers a rapid GAL4-dependent induction of galactose/melibiose regulon {GAL/MEL) gene transcription, and glucose represses this activation. We discovered that alterations in the physical state of the GAL4 protein correlate with activation and repression of the GAL/MEL genes. Using Western immunoblot assay, we observe two electrophoretic forms of GAL4 protein-GAL4i and GAL4n-in noninduced cells. In the absence of glucose, the addition of galactose to such cells results in the rapid appearance of a third and slowermigrating form, GAL4in, which differs from at least GAL4i by phosphorylation. CAI80-deletion cells that constitutively transcribe galactose-responsive genes due to the lack of the GAL80 protein, an antagonist of the GAL4 protein, exhibit GAL4ni without galactose addition. Addition of glucose, which results in rapid repression of galactose gene transcription, triggers a rapid elimination of GAL4in and an increase in GAL4u. Cycloheximide experiments provide evidence that the galactose-and glucose-triggered GAL4 protein mobility shifts are due to post-translational modification. GAL4ni is labeled with ppjphosphate in vivo; in vivo ^^S-labeled GAL4ni could be converted by phosphatase treatment in vitro to GAL4i. We present a model proposing that phosphorylation state changes in the GAL4 protein are key to modulating its activity.

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The design of a robust nonlinear feedback controller is analyzed for temperature control of continuous stirred tank reactors (CSTRs) which have strong nonlinearities and steady-state multiplicities. The present method treats the original nonlinear system as it is without transforming into an equivalent linear system. The controller is robust to modelling errors and random disturbances occurring in the system. The controller design is also analyzed for situation when the kinetics, activation energy and heat of reaction are unknown and also only limited state-variables measurements are available.

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Gal3p is an allosteric monomeric protein which activates the GAL genetic switch of Saccharomyces cerevisiae in response to galactose. Expression of constitutive mutant of Gal3p or over-expression of wild-type Gal3p activates the GAL switch in the absence of galactose.These data suggest that Gal3p exists as an ensemble of active and inactive conformations.Structural data has indicated that Gal3p exists in open (inactive) and closed (active) conformations. However, mutant of Gal3p that predominantly exists in inactive conformation and yet capable of responding to galactose has not been isolated. To understand the mechanism of allosteric transition, we have isolated a triple mutant of Gal3p with V273I, T404A and N450D substitutions which upon over-expression fails to activate the GAL switch on its own, but activates the switch in response to galactose. Over-expression of Gal3p mutants with single or double mutations in any of the three combinations failed to exhibit the behavior of the triple mutant. Molecular dynamics analysis of the wild-type and the triple mutant along with two previously reported constitutive mutants suggests that the wild-type Gal3p may also exist in super-open conformation. Further, our results suggest that the dynamics of residue F237 situated in the hydrophobic pocket located in the hinge region drives the transition between different conformations. Based on our study and what is known in human glucokinase, we suggest that the above mechanism could be a general theme in causing the allosteric transition.

Bioscientists such as geneticists and molecular biologists regularly demonstrate the integration of domain concepts and science inquiry practices/skills while explaining a natural phenomenon. The complexity of these concepts and skills becomes manifold at the tertiary undergraduate level and are known to be challenging for learners. They learn these in silos as part of theory classes, practical labs, and tutorial sessions while in an industry, they will be required to integrate and apply in a given authentic context. To support learners in this process, we have designed and developed Geneticus Investigatio (GI), a technology-enhanced learning (TEL) environment for scaffolding complex learning in the context of Mendelian genetics. GI facilitates this complex learning by the integration of domain concepts and science inquiry practices through inquiry-driven reflective learning experiences, which are interspersed with inquiry-based learning steps in an authentic context along with metacognitive reflection. In this paper, we present two cycles of iterative design, development, and evaluation of GI, based on the design-based research (DBR) approach. In the first DBR cycle, we identified the pedagogical design features and learning activities of GI based on an exploratory study with bio-science instructors for facilitating complex learning. We then report a pre-post classroom study (N = 37) in which we investigated the learning and perceptions of usability and usefulness of GI. The results indicate high learning gains after interacting with GI and learner perceptions that activities in GI help learn concepts and inquiry practices along with its integration. It is followed by the identification of interaction and other difficulties by the learner, which were triangulated with different data sources. It provided insights into the pedagogical and design changes required in GI. The revised version of GI was evaluated with a quasi-experimental classroom study (N = 121). The results indicate that the drawbacks of the previous version of GI were addressed. The main contributions of this research are a pedagogical design for facilitating complex learning and its implementation in the form of GI TEL environment.

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