Mehmet Sarikaya

Hsp70 and Hsp40 attenuate formation of spherical and annular polyglutamine oligomers by partitioning monomer

Protein conformational changes that result in misfolding, aggregation and amyloid fibril formation are a common feature of many neurodegenerative disorders. Studies with β-amyloid (Aβ), α-synuclein and other amyloid-forming proteins indicate that the assembly of misfolded protein conformers into fibrils is a complex process that may involve the population of metastable spherical and/or annular oligomeric assemblies. Here, we show by atomic force microscopy that a mutant huntingtin fragment with an expanded polyglutamine repeat forms spherical and annular oligomeric structures reminiscent of those formed by Aβ and α-synuclein. Notably, the molecular chaperones Hsp70 and Hsp40, which are protective in animal models of neurodegeneration, modulate polyglutamine aggregation reactions by partitioning monomeric conformations and disfavoring the accretion of spherical and annular oligomers.

Direct interaction of ligand–receptor pairs specifying stomatal patterning

Valves on the plant epidermis called stomata develop according to positional cues, which likely involve putative ligands (EPIDERMAL PATTERNING FACTORS [EPFs]) and putative receptors (ERECTA family receptor kinases and TOO MANY MOUTHS [TMM]) in Arabidopsis. Here we report the direct, robust, and saturable binding of bioactive EPF peptides to the ERECTA family. In contrast, TMM exhibits negligible binding to EPF1 but binding to EPF2. The ERECTA family forms receptor homomers in vivo. On the other hand, TMM associates with the ERECTA family but not with itself. While ERECTA family receptor kinases exhibit complex redundancy, blocking ERECTA and ERECTA-LIKE1 (ERL1) signaling confers specific insensitivity to EPF2 and EPF1, respectively. Our results place the ERECTA family as the primary receptors for EPFs with TMM as a signal modulator and establish EPF2-ERECTA and EPF1-ERL1 as ligand-receptor pairs specifying two steps of stomatal development: initiation and spacing divisions.

Biological Organization of Hydroxyapatite Crystallites into a Fibrous Continuum Toughens and Controls Anisotropy in Human Enamel

Enamel forms the outer surface of teeth, which are of complex shape and are loaded in a multitude of ways during function. Enamel has previously been assumed to be formed from discrete rods and to be markedly aniostropic, but marked anisotropy might be expected to lead to frequent fracture. Since frequent fracture is not observed, we measured enamel organization using histology, imaging, and fracture mechanics modalities, and compared enamel with crystalline hydroxyapatite (Hap), its major component. Enamel was approximately three times tougher than geologic Hap, demonstrating the critical importance of biological manufacturing. Only modest levels of enamel anisotropy were discerned; rather, our measurements suggest that enamel is a composite ceramic with the crystallites oriented in a complex three-dimensional continuum. Geologic apatite crystals are much harder than enamel, suggesting that inclusion of biological contaminants, such as protein, influences the properties of enamel. Based on our findings, we propose a new structural model.

Nano-mechanical properties profiles across dentin–enamel junction of human incisor teeth

Abstract Understanding how load is transferred from enamel to dentin and how the two tissues function as a single mechanical unit during mastication requires studies of micromechanics in relation to microstructure of the dentin–enamel junction (DEJ) zone. In this investigation, nano-hardness and elastic modulus of human incisor teeth were studied across the DEJ. It was found that, over a length scale of about 20 μm, there were decreasing trends in both hardness and elastic modulus across the DEJ zone profiling from enamel to dentin. Images obtained using atomic force microscopy from polished surfaces of cross-sectioned dental samples showed an interpenetrated microstructure of enamel and dentin at the DEJ zone. This result suggests that the nano-mechanical property profiles across the DEJ were due to a continuous variation in the ratios of relative amount of enamel and dentin. These characteristics of the DEJ zone could be significant for describing the structural and mechanical coupling of the two tissues. By increasing the contact area across the interface between the two hard tissues the stresses are dissipated reducing interfacial stress concentrations at the DEJ, thereby promoting effective load transfer from the hard (brittle) enamel to soft (tough) dentin.

Effect of Molecular Conformations on the Adsorption Behavior of Gold-Binding Peptides

Despite extensive recent reports on combinatorially selected inorganic-binding peptides and their bionanotechnological utility as synthesizers and molecular linkers, there is still only limited knowledge about the molecular mechanisms of peptide binding to solid surfaces. There is, therefore, much work that needs to be carried out in terms of both the fundamentals of solid-binding kinetics of peptides and the effects of peptide primary and secondary structures on their recognition and binding to solid materials. Here we discuss the effects of constraints imposed on FliTrx-selected gold-binding peptide molecular structures upon their quantitative gold-binding affinity. We first selected two novel gold-binding peptide (AuBP) sequences using a FliTrx random peptide display library. These were, then, synthesized in two different forms: cyclic (c), reproducing the original FliTrx gold-binding sequence as displayed on bacterial cells, and linear (l) dodecapeptide gold-binding sequences. All four gold-binding peptides were then analyzed for their adsorption behavior using surface plasmon resonance spectroscopy. The peptides exhibit a range of binding affinities to and adsorption kinetics on gold surfaces, with the equilibrium constant, Keq, varying from 2.5x10(6) to 13.5x10(6) M(-1). Both circular dichroism and molecular mechanics/energy minimization studies reveal that each of the four peptides has various degrees of random coil and polyproline type II molecular conformations in solution. We found that AuBP1 retained its molecular conformation in both the c- and l-forms, and this is reflected in having similar adsorption behavior. On the other hand, the c- and l-forms of AuBP2 have different molecular structures, leading to differences in their gold-binding affinities.

Rigid biological composite materials: Structural examples for biomimetic design

Biological hard tissues are composites of inorganics and biopolymers, and, therefore, represent hybrid systems. The inorganic components may be oxides (e.g., SiO2, Fe3O4), carbonates (e.g., CaCO3) sulfides (e.g., FeS, CdS), or others, mostly in crystalline forms but also occasionally in glassy forms. The biopolymer is often proteinaceous, but can also involve lipids and especially polysaccharides (e.g., chitin). These hybrid materials can be found in single celled organisms (such as bacteria and protozoa), invertebrates (such as mollusks), insects (such as beetles), and vertebrates (such as mammals). A common denominator of all hard tissues is that they are hierarchically structured from the nanometer scale to the microscale and the macroscale. It is these controlled structures that give biological hard tissues their unique and highly evolved functional properties. The engineering properties include mechanical, piezoelectric, optical, and magnetic. The hard tissues can be in the form of nanoparticles, spines, spicules, skeletons, and shells. The objective of this paper is to demonstrate mechanical aspects of some of these hard tissues, to discuss their structure-function relationships (with examples from the literature as well as from our research), and to reveal their potential utility in materials science and engineering applications.

Genetically engineered gold-binding polypeptides: structure prediction and molecular dynamics.

The biological control of inorganic crystal formation, morphology, and assembly is of interest to biologists and biotechnologists studying hard tissue growth and regeneration, as well as to materials scientists using biomimetic approaches for the control of inorganic material fabrication and assembly. Biomimetics requires an accurate understanding of natural mechanisms at the molecular level. Such understanding can be derived from the use of metal surfaces to study surface recognition by proteins together with combinatorial genetics techniques for the selection of suitable peptides. Polymerization of these peptides produces engineered polypeptides large enough to encode their own folding information with low structural complexity, while enhancing binding affinity to surfaces. The low complexity of such polypeptides can aid in analyses, leading to modeling and eventual manipulation of the structure of the folded polypeptide. This paper presents structure predictions for gold-binding protein sequences, originally selected by combinatorial techniques. Molecular dynamics simulations lasting 5 ns were carried out using solvated polypeptides at the gold surface to assess the dynamics of the binding process and the effects of surface topography on the specificity of protein binding.

A novel knowledge-based approach to design inorganic-binding peptides

MOTIVATION The discovery of solid-binding peptide sequences is accelerating along with their practical applications in biotechnology and materials sciences. A better understanding of the relationships between the peptide sequences and their binding affinities or specificities will enable further design of novel peptides with selected properties of interest both in engineering and medicine. RESULTS A bioinformatics approach was developed to classify peptides selected by in vivo techniques according to their inorganic solid-binding properties. Our approach performs all-against-all comparisons of experimentally selected peptides with short amino acid sequences that were categorized for their binding affinity and scores the alignments using sequence similarity scoring matrices. We generated novel scoring matrices that optimize the similarities within the strong-binding peptide sequences and the differences between the strong- and weak-binding peptide sequences. Using the scoring matrices thus generated, a given peptide is classified based on the sequence similarity to a set of experimentally selected peptides. We demonstrate the new approach by classifying experimentally characterized quartz-binding peptides and computationally designing new sequences with specific affinities. Experimental verifications of binding of these computationally designed peptides confirm our predictions with high accuracy. We further show that our approach is a general one and can be used to design new sequences that bind to a given inorganic solid with predictable and enhanced affinity.

Regulation of in vitro calcium phosphate mineralization by combinatorially selected hydroxyapatite-binding peptides.

We report selection and characterization of hydroxyapatite-binding heptapeptides from a peptide-phage library and demonstrate the effects of two peptides, with different binding affinities and structural properties, on the mineralization of calcium phosphate mineral. In vitro mineralization studies carried out using one strong- and one weak-binding peptide, HABP1 and HABP2, respectively, revealed that the former exhibited a drastic outcome on mineralization kinetics and particle morphology. Strong-binding peptide yielded significantly larger crystals, as observed by electron microscopy, in comparison to those formed in the presence of a weak-binding peptide or in the negative control. Molecular structural studies carried out by circular dichroism revealed that HABP1 and HABP2 differed in their secondary structure and conformational stability. The results indicate that sequence, structure, and molecular stability strongly influence the mineralization activity of these peptides. The implication of the research is that the combinatorially selected short-sequence peptides may be used in the restoration or regeneration of hard tissues through their control over of the formation of calcium phosphate biominerals.

Biomimetic model of a sponge-spicular optical fiber—mechanical properties and structure

Nanomechanical properties, nanohardness and elastic modulus, of an Antarctic sponge Rosella racovitzea were determined by using a vertical indentation system attached to an atomic force microscope. The Rosella spicules, known to have optical waveguide properties, are 10–20 cm long with a circular cross section of diameter 200–600 μm. The spicules are composed of 2–10-μm-thick layers of siliceous material that has no detectable crystallinity. Measurements through the thickness of the spicules indicated uniform properties regardless of layering. Both the elastic modulus and nanohardness values of the spicules are about half of that of either fused silica or commercial glass optical fibers. The fracture strength and fracture energy of the spicules, determined by 3-point bend tests, are several times those of silica rods of similar diameter. These sponge spicules are highly flexible and tough possibly because of their layered structure and hydrated nature of the silica. The spicules offer bioinspired lessons for potential biomimetic design of optical fibers with long-term durability that could potentially be fabricated at room temperature in aqueous solutions.