Biomedical subjects

Audun Bakk

Publications and source records attributed to Audun Bakk.

7 recordsLinked to original sources

PubMed2004

Transcriptional activation mechanisms of the PRM promoter of lambda phage.

We investigate the transcriptional activity associated with the P(RM) promoter of lambda phage. The probability for formation of a transcriptionally active (open) RNA polymerase-DNA complex is calculated by means of an equilibrium statistical-mechanical model. In particular, we study two different models of the transcriptional activation mechanism when the O(R)2 site is occupied by a CI dimer (typical for a lysogen) compared to the situation when O(R)2 is vacant: (1) transcription rate increases (wild-type mechanism) or (2) RNA polymerase becomes stronger (or weaker) bound to DNA (mutant mechanism). By applying experimental determined protein-DNA binding energies we show that these two mechanisms exhibit different characteristics when we study the activity versus CI concentration. We also show that our model may be fitted to in vivo activity data satisfactorily despite that the activated transcription rate is significantly reduced compared to the wild-type value. The model is consistent with experimental determined activities based upon mutations of CI and RNA polymerase.

Bacteriophage lambda↗

PubMed2004

Nonspecific binding of the OR repressors CI and Cro of bacteriophage lambda.

We estimate the Gibbs free energy for nonspecific binding (DeltaGNSB) to the Escherichia coli DNA for two regulatory proteins of the lambda phage, CI and Cro. By means of a statistical-mechanical approach, we calculate the cI and cro activities associated with the operator OR of an introduced lambda phage genome (prophage). In this statistical model we apply in vitro-measured binding free energies to fit in vivo experimental data for cI and cro activities, respectively, where DeltaGNSB is introduced as a free (fitting) parameter. Without nonspecific binding included in the model, the quality of the description is fairly poor, whereas data are nicely correlating with our model with nonspecific binding included over the entire data range. The obtained values of DeltaGNSB are -4.1+/-0.9 kcal/mol, for CI, and -4.2+/-0.8 kcal/mol, for Cro. In particular, in a lysogen (approximately 250 CI monomers per cell) we conclude that 86% of the total CI in the cell is nonspecifically bound, leaving on average around 10 CI dimers freely available in the E. coli cytoplasma. These findings corroborate the view that due to low free cellular particle numbers a dynamical analysis of genetic regulation at OR and comparable systems should include a stochastic component. In addition, we perform a stability analysis of the OR system in the presence of nonspecific binding.

Bacteriophage lambda↗

PubMed2004

In vivo non-specific binding of lambda CI and Cro repressors is significant.

We propose a thermodynamic model that includes the non-specific binding of the lambda phage regulatory proteins CI and Cro. By fitting the model to experimental in vivo data on activities of the two promoters P(RM) and P(R) versus concentration, we estimate the free energy upon non-specific binding to be -4.1+/-0.9 kcal/mol for CI and -4.2+/-0.8 kcal/mol for Cro. For concentrations >100 nM of CI or Cro, we find that >50% of these proteins are non-specifically bound. In particular, in a lysogen (approximately 250 CI monomeric equivalents per cell) nearly 90% of CI is non-specifically bound.

Bacteriophage lambda↗

PubMed2004

Sensitivity of OR in phage lambda.

We investigate the sensitivity of the right operator in bacteriophage lambda. In particular, the system is probed in the three different regulatory protein concentration-regimes: 1), lysogen (CI dominates); 2), during induction (CI and Cro at comparable concentrations); and 3), after induction (Cro dominates). Systematic perturbations of the protein-operator binding energies show in a lysogen that the activity (production rate) at promoter PRM is robust to variations, in contrast to PR, where the sensitivity is high. Both promoters, however, show large sensitivity in regimes 2 and 3. In all regimes we identify several suppressors, meaning that for a given large perturbation (+/-2 kcal/mol) of one binding energy, there exist compensating perturbation(s) that restore the wild-type activity.

Bacteriophage lambda↗

PubMed2004

Is it always possible to distinguish two- and three-state systems by evaluating the van't Hoff enthalpy?

Many small globular proteins are traditionally classified as thermodynamical two-state systems, i.e., the protein is either in the native, active state (folded) or in the denatured state (unfolded). We challenge this view and show that there may exist (protein) systems for which a van't Hoff analysis of experimental data cannot determine whether the system corresponds to two or three thermodynamical states when only temperatures in a narrow temperature region around the transition are considered. We generalize a widely employed two-state protein folding model to include a third, transition state. For this three-state system we systematically study the deviation of the calorimetric enthalpy (heat of transition) from the van't Hoff enthalpy, a measure of the two-stateness of a transition. We show that under certain conditions the heat capacity of the three-state system can be almost indistinguishable from the heat capacity for the two-state system over a broad temperature interval. The consequence may be that some three-state (or even more than three-states) systems have been misinterpreted as two-state systems when the conclusion is drawn solely upon the van't Hoff enthalpy. These findings are important not only for proteins, but also for the interpretation of thermodynamical systems in general.

Calorimetry↗

PubMed2002

Viscosity and transient electric birefringence study of clay colloidal aggregation.

We study a synthetic clay suspension of laponite at different particle and NaCl concentrations by measuring stationary shear viscosity and transient electrically induced birefringence (TEB). On one hand the viscosity data are consistent with the particles being spheres and the particles being associated with large amount bound water. On the other hand the viscosity data are also consistent with the particles being asymmetric, consistent with single laponite platelets associated with a very few monolayers of water. We analyze the TEB data by employing two different models of aggregate size (effective hydrodynamic radius) distribution: (1) bidisperse model and (2) log-normal distributed model. Both models fit, in the same manner, fairly well to the experimental TEB data and they indicate that the suspension consists of polydisperse particles. The models also appear to confirm that the aggregates increase in size vs increasing ionic strength. The smallest particles at low salt concentrations seem to be monomers and oligomers.

Journal Article↗

PubMed2002

Apolar and polar solvation thermodynamics related to the protein unfolding process.

Thermodynamics related to hydrated water upon protein unfolding is studied over a broad temperature range (5-125 degrees C). The hydration effect arising from the apolar interior is modeled as an increased number of hydrogen bonds between water molecules compared with bulk water. The corresponding contribution from the polar interior is modeled as a two-step process. First, the polar interior breaks hydrogen bonds in bulk water upon unfolding. Second, due to strong bonds between the polar surface and the nearest water molecules, we assume quantization using a simplified two-state picture. The heat capacity change upon hydration is compared with model compound data evaluated previously for 20 different proteins. We obtain good correspondence with the data for both the apolar and the polar interior. We note that the effective coupling constants for both models have small variations among the proteins we have investigated.

Biophysical Phenomena↗