Glycosylation May Reduce Protein Thermodynamic Stability by Inducing a Conformational Distortion.

Glycosylation plays not only a functional role but can also modify the biophysical properties of the modified protein. Usually, natural glycosylation results in protein stabilization; however, in vitro and in silico studies showed that sometimes glycosylation results in thermodynamic destabilization. Here, we applied coarse-grained and all-atom molecular dynamics simulations to understand the mechanism underlying the loss of stability of the MM1 protein by glycosylation.

Criteria for selecting PEGylation sites on proteins for higher thermodynamic and proteolytic stability.

PEGylation of protein side chains has been used for more than 30 years to enhance the pharmacokinetic properties of protein drugs. However, there are no structure- or sequence-based guidelines for selecting sites that provide optimal PEG-based pharmacokinetic enhancement with minimal losses to biological activity. We hypothesize that globally optimal PEGylation sites are characterized by the ability of the PEG oligomer to increase protein conformational stability; however, the current understanding of how PEG influences the conformational stability of proteins is incomplete.

The direction of protein entry into the proteasome determines the variety of products and depends on the force needed to unfold its two termini.

Poorly structured domains in proteins enhance their susceptibility to proteasomal degradation. To learn whether the presence of such a domain near either end of a protein determines its direction of entry into the proteasome, directional translocation was enforced on several proteasome substrates. Using archaeal PAN-20S complexes, mammalian 26S proteasomes, and cultured cells, we identified proteins that are degraded exclusively from either the C or N terminus and some showing no directional preference.

Nonnative interactions regulate folding and switching of myristoylated protein.

We present an integrated experimental and computational study of the molecular mechanisms by which myristoylation affects protein folding and function, which has been little characterized to date. Myristoylation, the covalent linkage of a hydrophobic C14 fatty acyl chain to the N-terminal glycine in a protein, is a common modification that plays a critical role in vital regulated cellular processes by undergoing reversible energetic and conformational switching. Coarse-grained folding simulations for the model pH-dependent actin- and membrane-binding protein hisactophilin reveal that nonnative hydrophobic interactions of the myristoyl with the protein as well as nonnative electrostatic interactions have a pronounced effect on folding rates and thermodynamic stability.

Communication: folding of glycosylated proteins under confinement.

Conjugating flexible polymers (such as oligosaccharides) to proteins or confining a protein in a restricted volume often increases protein thermal stability. In this communication, we investigate the interplay between conjugation and confinement which is not trivial as the magnitude and the mechanism of stabilization are different in each instance. Using coarse-grained computational approach the folding biophysics is studied when the protein is placed in a sphere of variable radius and is conjugated to 0-6 mono- or penta-saccharides.

Interaction of an antimicrobial peptide with membranes: experiments and simulations with NKCS.

We used Monte Carlo simulations and biophysical measurements to study the interaction of NKCS, a derivative of the antimicrobial peptide NK-2, with a 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE) membrane. The simulations showed that NKCS adsorbed on the membrane surface and the dominant conformation featured two amphipathic helices connected by a hinge region. We designed two mutants in the hinge to investigate the interplay between helicity and membrane affinity.

Folding of glycoproteins: toward understanding the biophysics of the glycosylation code.

Glycosylation is among the most common post-translational modifications that proteins undergo that may affect many of their activities. It may also modify the underlying energy landscape of glycoproteins in a way that their altered biophysical characteristics are linked to their bioactivity. Yet, the capability of glycosylation to modify thermodynamic and kinetic properties varies greatly between glycoproteins.

Effect of glycosylation on protein folding: a close look at thermodynamic stabilization.

Glycosylation is one of the most common posttranslational modifications to occur in protein biosynthesis, yet its effect on the thermodynamics and kinetics of proteins is poorly understood. A minimalist model based on the native protein topology, in which each amino acid and sugar ring was represented by a single bead, was used to study the effect of glycosylation on protein folding. We studied in silico the folding of 63 engineered SH3 domain variants that had been glycosylated with different numbers of conjugated polysaccharide chains at different sites on the protein's surface.

Interactions of cationic-hydrophobic peptides with lipid bilayers: a Monte Carlo simulation method.

We present a computational model of the interaction between hydrophobic cations, such as the antimicrobial peptide, Magainin2, and membranes that include anionic lipids. The peptide's amino acids were represented as two interaction sites: one corresponds to the backbone alpha-carbon and the other to the side chain. The membrane was represented as a hydrophobic profile, and its anionic nature was represented by a surface of smeared charges.

Has the code for protein translocation been broken?

Polypeptides chains are segregated by the translocon channel into secreted or membrane-inserted proteins. Recent reports claim that an in vivo system has been used to break the "amino acid code" used by translocons to make the determination of protein type (i.e.

Monte Carlo studies of folding, dynamics, and stability in alpha-helices.

Folding simulations of polyalanine peptides were carried out using an off-lattice Monte Carlo simulation technique. The peptide was represented as a chain of residues, each of which contains two interaction sites: one corresponding to the C(alpha) atom and the other to the side chain. A statistical potential was used to describe the interaction between these sites.

Interactions of hydrophobic peptides with lipid bilayers: Monte Carlo simulations with M2delta.

We introduce here a novel Monte Carlo simulation method for studying the interactions of hydrophobic peptides with lipid membranes. Each of the peptide's amino acids is represented as two interaction sites: one corresponding to the backbone alpha-carbon and the other to the side chain, with the membrane represented as a hydrophobic profile. Peptide conformations and locations in the membrane and changes in the membrane width are sampled using the Metropolis criterion, taking into account the underlying energetics.

Long-range effects on the binding of the influenza HA to receptors are mediated by changes in the stability of a metastable HA conformation.

X-ray studies show that influenza hemagglutinin (HA) forms an elongated structure connecting the influenza virus at one end to cell-surface receptors at the other. At neutral pH, the 20 N-terminal residues of HA2-referred to as the fusion peptide-are buried in a hydrophobic pocket, about 100 A away from the receptor-binding site, and thus seem unlikely to affect HA binding to the receptor. To test this assumption, we mutated residues in the fusion peptide, heterologically expressed the mutated proteins in COS7 cells, and examined their ability to bind fluorescently labeled red blood cells (RBCs).