Ordered-domain unfolding of thermophilic isolated ß subunit ATP synthase.

The flexibility of the ATP synthase's ß subunit promotes its role in the ATP synthase rotational mechanism, but its domains stability remains unknown. A reversible thermal unfolding of the isolated ß subunit (Tß) of the ATP synthase from Bacillus thermophilus PS3, tracked through circular dichroism and molecular dynamics, indicated that Tß shape transits from an ellipsoid to a molten globule through an ordered unfolding of its domains, preserving the ß-sheet residual structure at high temperature. We determined that part of the stability origin of Tß is due to a transversal hydrophobic array that crosses the ß-barrel formed at the N-terminal domain and the Rossman fold of the nucleotide-binding domain (NBD), while the helix bundle of the C-terminal domain is the less stable due to the lack of hydrophobic residues, and thus the more flexible to trigger the rotational mechanism of the ATP synthase.

Improving coarse-grained models of protein folding through weighting of polar-polar/hydrophobic-hydrophobic interactions into crowded spaces.

Herein were tested 7 hydrophobic-polar sequences in two types of 2D-square space lattices, homogeneous and correlated, the latter simulating molecular crowding included as a geometric boundary restriction. Optimization of 2D structures was carried out using a variant of Dill's model, inspired by convex function, taking into account both hydrophobic (Dill's model) and polar interactions, including more structural information to reach better folding solutions. While using correlated networks, degrees of freedom in the folding of sequences were limited; as a result in all cases, more successful structural trials were found in comparison to a homogeneous lattice. The majority of employed sequences were designed by our workgroup, two of them were folded with other approaches, and another is a modified version of a previous sequence, initial forms of the other two have been employed but without taking into account polar-polar contributions. Three of them are newly proposed, intended to test the conjoint hydrophobic-hydrophobic and polar-polar contributions in crowded spaces. One sequence turned out to be the most difficult of the seven folded, this perhaps due to intrinsic (i) degrees of freedom and (ii) motifs of the expected 2D HP structure. Meanwhile two-sequence, although optimal folding was not achieved for neither of the two approaches, folding with correlated network approach not only produced better results than homogeneous space, but for them the best values found with crowding were very close to the expected optimal fitness. In general, five sequences were better folded with medium lattice units for correlated media; instead, another two sequences were better folded with a bit larger degree of lattice unit, revealing that depending on the degrees of freedom and particular folding, motifs in each sequence would require tuned crowding to achieve better folding. Therefore, the main goal herein was to obtain a modified 2D HP lattice model to mimic folding of proteins or secondary structures, like ß-sheets, taking into account both hydrophobic-hydrophobic and polar-polar interactions, and fold them in a crowded environment. This simple but enough construction would be conducted to determine the needed information to fold sequences in a sort of a minimal but complete heuristic model. Finally, we claim that all folded sequences into crowded spaces achieve better results than homogeneous ones.

Nanostructured oleic acid/polysorbate 80 emulsions with diminished toxicity in NL-20 cell line: Insights of potential drug carriers.

Nanoemulsions (NE) are nowadays required drug nanocarriers. We have selected i) oleic acid (OA) as oil (O), ii) polysorbate 80 (PS80) as surfactant (S), and iii) water (W) in a prototype NE. Our best formulation had O:S ratio [OA]/[PS80] = 0.0708/0.0382 = 1.85 [mol·L-1], implying 1.85 parts of OA covered/stabilized by 1 part of PS80, giving 71.86 nm and 0.42 polydispersity index (PDI) in NE, determined by DLS and TEM. These nanosystems stored at room temperature/darkness stabilized up to 12 months (measured by DLS and TEM) maintaining very similar particle sizes and sometimes decreasing PDI. NE stability was determined by DSC, evidencing reversibility upon heating from 25 to 100 °C, increasing to 125 °C (sealed systems) produced more attenuated heating profiles in second and third cycles, compared with first, indicating partial but enough stability for storage means. NE cytotoxicity tests were conducted on immortalized normal lung epithelial cells (NL-20), as reference. The results show 50 % inhibitory concentrations (IC50,µM) of 1100, OA, and 2.6, PS80. The IC50 was 20.5, PS80 (PS80@NE) and 37.9, OA (OA@NE) clearly indicating that components changed their toxicities upon nanostructuring, OA exhibited 30-fold increase (IC50(OA) 1100.0→37.9) while PS80, decreased 7.9-fold (IC50(PS80) 2.6→20.5). PS80 is the most toxic component but when is included in PS80@NE, less toxic nanocarriers were generated.

In silico minimalist approach to study 2D HP protein folding into an inhomogeneous space mimicking osmolyte effect: First trial in the search of foldameric backbones.

We have employed our bioinformatics workbench, named Evolution, a Multi-Agent System based architecture with lattice-bead-models, evolutionary-algorithms, and correlated-networks as inhomogeneous spaces, with different correlation lengths, mimicking osmolyte effect (molecular crowding), to in silico survey protein folding. Resolution is with hydrophobic-polar (H-P) sequences in inhomogeneous 2D square lattices, since general biophysicochemical trends consider i) that the backbone is one of the major components responsible for protein folding and ii) osmolyte effect plays an important role to better folding kinetics and reach deeper optima. We have designed foldamers, as square n × n (n = 3, 4, 5, 6) arrays of hydrophobic cores stabilized by H⋯H contacts, attached through short PP (P2) or long PPPP (P4) loops, giving rise to 8 sequences (S1 to S8) with known optimal scores. Designed sequences were folded into different inhomogeneous spaces and indeed crowded media induced deeper optima, being crowding necessary to best fold, but the space should be enough constrained to induce folding without banning chain movement. The constrained space plays an important role to reach the optimal structure, depending on designed foldamer sequence size, for an optimal correlation length, implying that media affects the folding pathways as happens in real systems. Designed structures were found, moreover, they undergo to degenerated states, both folding states could survey considering i) backbone information and ii) osmolyte effect. In nature, the proteins fold in different structures aiming to reach a global minimum, but a local minimum could be enough to the protein to be functional or dysfunctional.

Photophysics of a cis axially disubstituted macrocycle: rapid intersystem crossing in a tin(IV) phthalocyanine with a half-domed geometry.

We have studied the photophysical properties of a tin(IV) phthalocyanine which coordinates two myristate groups through their carboxylate functionalities in a cis disposition at the tin center. Such a coordination mode, anisobidentate through the same side of the macrocycle, makes this phthalocyanine acquire a capped or half-domed shape. This bis myristate tin(IV) molecule shows an intersystem crossing channel which populates the triplet manifold with high efficiency and with a time constant of 300 ps, about an order of magnitude faster than planar phthalocyanines, including some previously reported tin(IV) phthalocyanines. For comparison purposes, we also include the description of a planar silicon(IV) phthalocyanine that keeps the more common stereochemistry, of trans type, with the same axial myristate groups. The characterization of these systems included steady state and time-resolved spectroscopy through femtosecond fluorescence up-conversion and transient absorption. We also studied the initial S(n) → S(1) internal conversion dynamics when these compounds are excited to upper states with 387.5 nm light. In addition, we include measurements of the rate for singlet oxygen production through the formation of an ESR-active adduct in aerated solutions. Such measurements indicate that, associated to its photophysics, the tin(IV) phthalocyanine produces (1)O(2) with an efficiency significantly larger than the silicon(IV) counterpart, making it an interesting option for sensitization applications. Finally, we performed excited state calculations at the TD-DFT level which describe the effects of the reduced symmetry together with the state ordering and indicate the presence of near dark intermediate states between the Q and B transitions for both of these macrocycles.