Biomolecules2013, 3(4), 986-996; doi:10.3390/biom3040986 - published online 6 December 2013 Show/Hide Abstract
Abstract: Charge-pair interactions between acidic and basic residues on the surface of collagen can promote stability as well as control specificity of molecular recognition. Heterotrimeric collagen peptides have been engineered de novo using either rational or computational methods, which in both cases optimize networks of favorable charge-pair interactions in the target structure. Less understood is the role of electrostatic repulsion between groups of like charge in destabilizing structure or directing molecular recognition. To study this, we apply a “charge crowding” approach, where repulsive interactions between multiple aspartate side chains are found to destabilize the homotrimer states in triple helical peptide system and can be utilized to promote the formation of heterotrimers. Neutralizing surface charge by increasing salt concentration or decreasing pH can enhance homotrimer stability, confirming the role of charge crowding on the destabilization of homotrimers via electrostatic repulsion. Charge crowding may be used in conjunction with other approaches to create specific collagen heterotrimers.
Biomolecules2013, 3(4), 967-985; doi:10.3390/biom3040967 - published online 18 November 2013 Show/Hide Abstract
Abstract: Euplotes nobilii and Euplotes raikovi are phylogenetically closely allied species of marine ciliates, living in polar and temperate waters, respectively. Their evolutional relation and the sharply different temperatures of their natural environments make them ideal organisms to investigate thermal-adaptation. We perform a comparative study of the thermal unfolding of disulfide-rich protein pheromones produced by these ciliates. Recent circular dichroism (CD) measurements have shown that the two psychrophilic (E. nobilii) and mesophilic (E. raikovi) protein families are characterized by very different melting temperatures, despite their close structural homology. The enhanced thermal stability of the E. raikovi pheromones is realized notwithstanding the fact that these proteins form, as a rule, a smaller number of disulfide bonds. We perform Monte Carlo (MC) simulations in a structure-based coarse-grained (CG) model to show that the higher stability of the E. raikovi pheromones is due to the lower locality of the disulfide bonds, which yields a lower entropy increase in the unfolding process. Our study suggests that the higher stability of the mesophilic E. raikovi phermones is not mainly due to the presence of a strongly hydrophobic core, as it was proposed in the literature. In addition, we argue that the molecular adaptation of these ciliates may have occurred from cold to warm, and not from warm to cold. To provide a testable prediction, we identify a point-mutation of an E. nobilii pheromone that should lead to an unfolding temperature typical of that of E. raikovi pheromones.
Biomolecules2013, 3(4), 943-966; doi:10.3390/biom3040943 - published online 11 November 2013 Show/Hide Abstract
Abstract: In eukaryotic cells, mitochondria are self-reproducing organelles with their own DNA and they play a central role in adenosine triphosphate (ATP) synthesis by respiration. Increasing evidence indicates that mitochondria also have critical and multiple functions in the initiation of cell differentiation, cell-type determination, cell movement, and pattern formation. This has been most strikingly realized in development of the cellular slime mold Dictyostelium. For example, the expression of the mitochondrial ribosomal protein S4 (mt-rps4) gene is required for the initial differentiation. The Dictyostelium homologue (Dd-TRAP1) of TRAP-1 (tumor necrosis receptor-associated protein 1), a mitochondrial molecular chaperone belonging to the Hsp90 family, allows the prompt transition of cells from growth to differentiation through a novel prestarvation factor (PSF-3) in growth medium. Moreover, a cell-type-specific organelle named a prespore-specific vacuole (PSV) is constructed by mitochondrial transformation with the help of the Golgi complex. Mitochondria are also closely involved in a variety of cellular activities including CN-resistant respiration and apoptosis. These mitochondrial functions are reviewed in this article, with special emphasis on the regulation of Dictyostelium development.
Biomolecules2013, 3(4), 923-942; doi:10.3390/biom3040923 - published online 8 November 2013 Show/Hide Abstract
Abstract: Proteolytic enzymes (also termed peptidases, proteases and proteinases) are capable of hydrolyzing peptide bonds in proteins. They can be found in all living organisms, from viruses to animals and humans. Proteolytic enzymes have great medical and pharmaceutical importance due to their key role in biological processes and in the life-cycle of many pathogens. Proteases are extensively applied enzymes in several sectors of industry and biotechnology, furthermore, numerous research applications require their use, including production of Klenow fragments, peptide synthesis, digestion of unwanted proteins during nucleic acid purification, cell culturing and tissue dissociation, preparation of recombinant antibody fragments for research, diagnostics and therapy, exploration of the structure-function relationships by structural studies, removal of affinity tags from fusion proteins in recombinant protein techniques, peptide sequencing and proteolytic digestion of proteins in proteomics. The aim of this paper is to review the molecular biological aspects of proteolytic enzymes and summarize their applications in the life sciences.
Biomolecules2013, 3(4), 905-922; doi:10.3390/biom3040905 - published online 6 November 2013 Show/Hide Abstract
Abstract: Understanding chemokine interactions with glycosaminoglycans (GAG) is critical as these interactions have been linked to a number of inflammatory medical conditions, such as arthritis and asthma. To better characterize in vivo protein function, comprehensive knowledge of multimeric species, formed by chemokines under native conditions, is necessary. Herein is the first report of a tetrameric assembly of the human chemokine CCL11, which was shown bound to the GAG Arixtra™. Isothermal titration calorimetry data indicated that CCL11 interacts with Arixtra, and ion mobility mass spectrometry (IM-MS) was used to identify ions corresponding to the CCL11 tetrameric species bound to Arixtra. Collisional cross sections (CCS) of the CCL11 tetramer-Arixtra noncovalent complex were compared to theoretical CCS values calculated using a preliminary structure of the complex deduced using X-ray crystallography. Experimental CCS values were in agreement with theoretical values, strengthening the IM-MS evidence for the formation of the noncovalent complex. Tandem mass spectrometry data of the complex indicated that the tetramer-GAG complex dissociates into a monomer and a trimer-GAG species, suggesting that two CC-like dimers are bridged by Arixtra. As development of chemokine inhibitors is of utmost importance to treatment of medical inflammatory conditions, these results provide vital insights into chemokine-GAG interactions.
Biomolecules2013, 3(4), 889-904; doi:10.3390/biom3040889 - published online 1 November 2013 Show/Hide Abstract
Abstract: HIV-1 reverse transcriptase (RT) plays a central role in HIV infection. Current United States Federal Drug Administration (USFDA)-approved antiretroviral therapies can include one of five approved non-nucleoside RT inhibitors (NNRTIs), which are potent inhibitors of RT activity. Despite their crucial clinical role in treating and preventing HIV-1 infection, their mechanism of action remains elusive. In this review, we introduce RT and highlight major advances from experimental and computational biophysical experiments toward an understanding of RT function and the inhibitory mechanism(s) of NNRTIs.