Commitment to folded and aggregated states occurs late in interleukin-1 beta folding.

A point mutation, lysine 97 to isoleucine, in the all-beta cytokine interleukin-1 beta (IL-1 beta) exhibits an increased propensity to form inclusion bodies in vivo and aggregates in vitro. In an effort to better understand the aggregation reaction and determine when intervention may allow rescue of protein from aggregation during renaturation, we developed a novel application of mass spectrometry using isotopic labeling to determine the step(s) at which K97I commits to either the native or aggregated state. Interestingly, despite the early formation of a folding intermediate ensemble at an observed rate lambda(2) of 4.

Aggregation events occur prior to stable intermediate formation during refolding of interleukin 1beta.

A point mutation, lysine 97 --> isoleucine (K97I), in a surface loop in the beta-sheet protein interleukin 1beta (IL-1beta), exhibits increased levels of inclusion body (IB) formation relative to the wild-type protein (WT) when expressed in Escherichia coli. Despite the common observation that less stable proteins are often found in IBs, K97I is more stable than WT. We examined the folding pathway of the mutant and wild-type proteins at pH 6.

Conformation-invariant structures of the alpha1beta1 human hemoglobin dimer.

Analysis of the conformational differences between the oxy and deoxy forms of hemoglobin is complicated by shifting coordinate systems and correlated motions between different parts of the molecule. Methods independent of any frame of reference were used to study the differences in structure between the oxy and deoxy forms of the human hemoglobin alphabeta dimer. Differences between the deoxy and oxy dimer structures can be characterized as rearrangements of 15 substructures persisting between the two conformations.

A gel as an array of channels.

We consider the theory of charged point molecules ('probes') being pulled by an electric field through a two-dimensional net of channels that represents a piece of gel. Associated with the position in the net is a free energy of interaction between the probe and the net; this free energy fluctuates randomly with the position of the probe in the net. The free energy is intended to represent weak interactions between the probe and the gel, such as entropy associated with the restriction of the freedom of motion of the probe by the gel, or electrostatic interactions between the probe and charges fixed to the gel.

Rigid domains in proteins: an algorithmic approach to their identification.

A rigid domain, defined here as a tertiary structure common to two or more different protein conformations, can be identified numerically from atomic coordinates by finding sets of residues, one in each conformation, such that the distance between any two residues within the set belonging to one conformation is the same as the distance between the two structurally equivalent residues within the set belonging to any other conformation. The distance between two residues is taken to be the distance between their respective alpha carbon atoms. With the methods of this paper we have found in the deoxy and oxy conformations of the human hemoglobin alpha 1 beta 1 dimer a rigid domain closely related to that previously identified by Baldwin and Chothia (J.

The degradation of T7 DNA in converging flow.

The flow-induced degradation of T7 DNA (Molecular Size = 40 Kbp) was studied in a flow device that generates converging flow rather than simple shear flow. We discovered that the sizes of the degradation products were very broadly distributed, covering the range from 10 Kbp to 36 Kbp. An explanation for the broadness of the distribution is given based on a computer simulation of the experiment.

Understanding the anomalous electrophoresis of bent DNA molecules: a reptation model.

In polyacrylamide gel electrophoresis, the retardation of DNA molecules containing regions of intrinsic curvature can be explained by a novel reptation model that includes the elastic free energy of the DNA chain. Computer simulations based on this model give results that reproduce the dependence of anomalous mobility on gel concentration, which is quantified by new experimental data on the mobilities of circularly permuted isomers of kinetoplast DNA fragments. Fitting of the data required allowing for the elasticity of the gel.

Separations of open-circular DNA using pulsed-field electrophoresis.

The effect of high electric fields on the gel-electrophoretic mobility of open-circular DNA in agarose differs dramatically from that on linear molecules of the same molecular weight. At high fields, sufficiently large circular forms are prevented from migrating into the gel whereas linear molecules and smaller circular DNAs migrate normally. This effect is strongly field dependent, affecting circular molecules of decreasing size with increasing field strength.

Counter-ion condensation and system dimensionality.

Condensation of the counter-ions around a highly charged infinitely long cylindrical molecule, such as DNA, can be described in detail in terms of the solutions of the Poisson-Boltzmann (Gouy-Chapman) equation. By using the Alfrey-Berg-Morawetz (1951) solution of this equation one can show that a certain fraction of the counter-ions remain within finite distances of the poly-ion even when the volume of the system is expanded indefinitely; these ions can be appropriately called "condensed". The fraction of the macromolecule's charge represented by these ions is just 1-1/xi, where xi is the linear charge-density parameter of the macromolecule; this is also the value given by Manning's theory.

A rhelogical separator for very large DNA molecules.

We present a rheological separation method for DNA molecules in which their deformability is used to advantage. This is the "radial migration method"; here we present experimental verification of the principle, theory having been reported elsewhere. The main conclusions are: (1) the theory is reasonably good; (2) radial migration is highly sensitive to the molecular weight, as predicted, and (3) intact T2 DNA (1.

Molecular weight of T2 NaDNA from viscoelasticity.

The viscoelastic properties of T2 DNA solutions are used to determine the NaDNA molecular weight in four independent ways from the theory of the beads-springs model. The four molecular weights are 131.9, 132.