Assembly of lipoprotein particles revealed by coarse-grained molecular dynamics simulations

High-density lipoproteins (HDL) function as cholesterol transporters, facilitating the removal of excess cholesterol from the body. Due to the heterogeneity of native HDL particles (both in size and shape), the details on how these protein-lipid particles form and the structure they assume in their lipid-associated states are not well characterized. We report here a study of the self-assembly of discoidal HDL particles using coarse-grained (CG) molecular dynamics. The microsecond simulations reveal the self-assembly of HDL particles from disordered protein-lipid complexes to form structures containing many of the features of the generally accepted double-belt model for discoidal HDL particles. HDL assembly is found to proceed in two broad steps, aggregation of proteins and lipids driven by the hydrophobic effect which occurs on a approximately 1 micros time scale, followed by the optimization of the protein structure driven by increasingly specific protein-protein interactions.     

Folding transition into a loosely collapsed state in plasmid DNA as revealed by single-molecule observation

The conformational transition of a plasmid DNA, pGEG.GL3 (12.5 kbp, circular), induced by spermine(4+) was studied through the observation of individual DNA by fluorescence microscopy. We deduced the change in the hydrodynamic radius R(H) from an analysis of the Brownian motion of single DNA molecules. R(H) decreases in a continuous manner with an increase in spermine(4+), in contrast to the large discrete on/off change for long linear DNA. Just after the transition to the collapsed state, a small number of DNA molecules tend to form an assembly, which disperses in the bulk solution without precipitation.     

Differential biological role of CD3 chains revealed by human immunodeficiencies

The biological role in vivo of the homologous CD3gamma and delta invariant chains within the human TCR/CD3 complex is a matter of debate, as murine models do not recapitulate human immunodeficiencies. We have characterized, in a Turkish family, two new patients with complete CD3gamma deficiency and SCID symptoms and compared them with three CD3gamma-deficient individuals belonging to two families from Turkey and Spain. All tested patients shared similar immunological features such as a partial TCR/CD3 expression defect, mild alphabeta and gammadelta T lymphocytopenia, poor in vitro proliferative responses to Ags and mitogens at diagnosis, and very low TCR rearrangement excision circles and CD45RA(+) alphabeta T cells. However, intrafamilial and interfamilial clinical variability was observed in patients carrying the same CD3G mutations. Two reached the second or third decade in healthy conditions, whereas the other three showed lethal SCID features with enteropathy early in life. In contrast, all reported human complete CD3delta (or CD3epsilon) deficiencies are in infants with life-threatening SCID and very severe alphabeta and gammadelta T lymphocytopenia. Thus, the peripheral T lymphocyte pool was comparatively well preserved in human CD3gamma deficiencies despite poor thymus output or clinical outcome. We propose a CD3delta > CD3gamma hierarchy for the relative impact of their absence on the signaling for T cell production in humans.     

Structure and interactions of RecA: plasticity revealed by molecular dynamics simulations

Eleven independent simulations, each involving three consecutive molecules in the RecA filament, carried out on the protein from Mycobacterium tuberculosis, Mycobacterium smegmatis and Escherichia coli and their Adenosine triphosphate (ATP) complexes, provide valuable information which is complementary to that obtained from crystal structures, in addition to confirming the robust common structural framework within which RecA molecules from different eubacteria function. Functionally important loops, which are largely disordered in crystal structures, appear to adopt in each simulation subsets of conformations from larger ensembles. The simulations indicate the possibility of additional interactions involving the P-loop which remains largely invariant. The phosphate tail of the ATP is firmly anchored on the loop while the nucleoside moiety exhibits substantial structural variability. The most important consequence of ATP binding is the movement of the ‘switch’ residue. The relevant simulations indicate the feasibility of a second nucleotide binding site, but the pathway between adjacent molecules in the filament involving the two nucleotide binding sites appears to be possible only in the mycobacterial proteins.     

Structure of spheroidal HDL particles revealed by combined atomistic and coarse-grained simulations

Spheroidal high-density lipoprotein (HDL) particles circulating in the blood are formed through an enzymatic process activated by apoA-I, leading to the esterification of cholesterol, which creates a hydrophobic core of cholesteryl ester molecules in the middle of the discoidal phospholipid bilayer. In this study, we investigated the conformation of apoA-I in model spheroidal HDL (ms-HDL) particles using both atomistic and coarse-grained molecular dynamics simulations, which are found to provide consistent results for all HDL properties we studied. The observed small contribution of cholesteryl oleate molecules to the solvent-accessible surface area of the entire ms-HDL particle indicates that palmitoyloleoylphosphatidylcholines and apoA-I molecules cover the hydrophobic core comprised of cholesteryl esters particularly well. The ms-HDL particles are found to form a prolate ellipsoidal shape, with sizes consistent with experimental results. Large rigid domains and low mobility of the protein are seen in all the simulations. Additionally, the average number of contacts of cholesteryl ester molecules with apoA-I residues indicates that cholesteryl esters interact with protein residues mainly through their cholesterol moiety. We propose that the interaction of annular cholesteryl oleate molecules contributes to apoA-I rigidity stabilizing and regulating the structure and function of the ms-HDL particle.     

Structural instability of the prion protein upon M205S/R mutations revealed by molecular dynamics simulations

The point mutations M205S and M205R have been demonstrated to severely disturb the folding and maturation process of the cellular prion protein (PrP(C)). These disturbances have been interpreted as consequences of mutation-induced structural changes in PrP, which are suggested to involve helix 1 and its attachment to helix 3, because the mutated residue M205 of helix 3 is located at the interface of these two helices. Furthermore, current models of the prion protein scrapie (PrP(Sc)), which is the pathogenic isoform of PrP(C) in prion diseases, imply that helix 1 disappears during refolding of PrP(C) into PrP(Sc). Based on molecular-dynamics simulations of wild-type and mutant PrP(C) in aqueous solution, we show here that the native PrP(C) structure becomes strongly distorted within a few nanoseconds, once the point mutations M205S and M205R have been applied. In the case of M205R, this distortion is characterized by a motion of helix 1 away from the hydrophobic core into the aqueous environment and a subsequent structural decay. Together with experimental evidence on model peptides, this decay suggests that the hydrophobic attachment of helix 1 to helix 3 at M205 is required for its correct folding into its stable native structure.     

Conformational behavior of polyethylenimine derivatives revealed by fluorescence spectroscopy

Conformation behavior of polyethylenimine has been examined by studies of the fluorescence characteristics of derivatives of the polymer containing pyrenyl ligands. Excimer formation within the macromolecular matrix serves as a sensitive probe of group proximities. The experimental observations combined with nearest neighbor analyses lead to quantitative assessments of the extent of interaction between polymer side chains.     

Interaction of diverse voltage sensor homologs with lipid bilayers revealed by self-assembly simulations

Homologous pairing and braiding (supercoiling) have crucial effects on genome organization, maintenance, and evolution. Generally, the pairing and braiding processes are discussed in different contexts, independently of each other. However, analysis of electrostatic interactions between DNA double helices suggests that in some situations these processes may be related. Here we present a theory of DNA braiding that accounts for the elastic energy of DNA double helices as well as for the chiral nature of the discrete helical patterns of DNA charges. This theory shows that DNA braiding may be affected, stabilized, or even driven by chiral electrostatic interactions. For example, electrostatically driven braiding may explain the surprising recent observation of stable pairing of homologous double-stranded DNA in solutions containing only monovalent salt. Electrostatic stabilization of left-handed braids may stand behind the chiral selectivity of type II topoisomerases and positive plasmid supercoiling in hyperthermophilic bacteria and archea.     

Differential helix propensity of small apolar side chains studied by molecular dynamics simulations.

A series of oligoalanine molecules with single amino acid replacements in the middle of the chain has been studied by molecular dynamics simulations. Differences in stability of the alpha-helix (as free energies delta delta G degrees) were estimated for the following series of residues: alpha-aminoisobutyric acid, alanine, alpha-amino-n-butyric acid, valine, glycine, D-alanine, t-leucine (= alpha-amino-beta,beta-dimethyl-n- butyric acid), and proline, arranged here in decreasing order of helix-forming potential. (The results for proline and valine had been reported earlier.) No experimental results were available for alpha-amino-n-butyric acid, D-alanine, and t-leucine at the time these calculations were done. The values of delta delta G degrees, including the three predictions, are in striking agreement with recent experimental results. A combination of free dynamics, dynamics with forced conformational change, and dynamics with forced molecular replacement was used. Conformational distributions were calculated for the peptide backbone of the dipeptides and, where appropriate, for the side chains of the dipeptide and the alpha-helix. The results demonstrate an unexpected level of accuracy for the all-atom model used to represent atomic interactions in the simulations. The simulations permit a detailed analysis of different factors responsible for conformational preferences and differences in stability. These conclusions drawn from this analysis agree with accepted qualitative explanations and allow these explanations to be quantitated to an extent not heretofore possible.