Design of net-charged abc-type collagen heterotrimers.
Ontology highlight
ABSTRACT:
SUBMITTER: Parmar AS
PROVIDER: S-EPMC3874424 | biostudies-literature | 2014 Feb
REPOSITORIES: biostudies-literature
Publications
Design of net-charged abc-type collagen heterotrimers.
Parmar Avanish S AS Zahid Sohail S Belure Sandeep V SV Young Robert R Hasan Nida N Nanda Vikas V
Journal of structural biology 20130418 2
Net-negatively-charged heterospecific A:B:C collagen peptide heterotrimers were designed using an automated computational approach. The design algorithm considers both target stability and the energy gap between the target states and misfolded competing states. Structural characterization indicates the net-negative charge balance on the new designs enhances the specificity of the target state at the expense of its stability. ...[more]
Similar Datasets
Project description:Normal type I collagen is a heterotrimer of two alpha1(I) and one alpha2(I) chains, but various genetic and environmental factors result in synthesis of homotrimers that consist of three alpha1(I) chains. The homotrimers completely replace the heterotrimers only in rare recessive disorders. In the general population, they may compose just a small fraction of type I collagen. Nevertheless, they may play a significant role in pathology; for example, synthesis of 10-15% homotrimers due to a polymorphism in the alpha1(I) gene may contribute to osteoporosis. Homotrimer triple helices have different stability and less efficient fibrillogenesis than heterotrimers. Their fibrils have different mechanical properties. However, very little is known about their molecular interactions and fibrillogenesis in mixtures with normal heterotrimers. Here we studied the kinetics and thermodynamics of fibril formation in such mixtures by combining traditional approaches with 3D confocal imaging of fibrils, in which homo- and heterotrimers were labeled with different fluorescent colors. In a mixture, following a temperature jump from 4 to 32 degrees C, we observed a rapid increase in turbidity most likely caused by formation of homotrimer aggregates. The aggregates promoted nucleation of homotrimer fibrils that served as seeds for mixed and heterotrimer fibrils. The separation of colors in confocal images indicated segregation of homo- and heterotrimers at a subfibrillar level throughout the process. The fibril color patterns continued to change slowly after the fibrillogenesis appeared to be complete, due to dissociation and reassociation of the pepsin-treated homo- and heterotrimers, but this remixing did not significantly reduce the segregation even after several days. Independent homo- and heterotrimer solubility measurements in mixtures confirmed that the subfibrillar segregation was an equilibrium property of intermolecular interactions and not just a kinetic phenomenon. We argue that the subfibrillar segregation may exacerbate effects of a small fraction of alpha1(I) homotrimers on formation, properties, and remodeling of collagen fibers.
| S-EPMC2839200 | biostudies-literature
Project description:We sought to computationally design model collagen peptides that specifically associate as heterotrimers. Computational design has been successfully applied to the creation of new protein folds and functions. Despite the high abundance of collagen and its key role in numerous biological processes, fibrous proteins have received little attention as computational design targets. Collagens are composed of three polypeptide chains that wind into triple helices. We developed a discrete computational model to design heterotrimer-forming collagen-like peptides. Stability and specificity of oligomerization were concurrently targeted using a combined positive and negative design approach. The sequences of three 30-residue peptides, A, B, and C, were optimized to favor charge-pair interactions in an ABC heterotrimer, while disfavoring the 26 competing oligomers (i.e., AAA, ABB, BCA). Peptides were synthesized and characterized for thermal stability and triple-helical structure by circular dichroism and NMR. A unique A:B:C-type species was not achieved. Negative design was partially successful, with only A + B and B + C competing mixtures formed. Analysis of computed versus experimental stabilities helps to clarify the role of electrostatics and secondary-structure propensities determining collagen stability and to provide important insight into how subsequent designs can be improved.
| S-EPMC2853261 | biostudies-literature
Project description:Collagen mimetic peptides (CMPs) provide critical insight into the assembly, stability, and structure of the triple helical collagen protein. The majority of natural fibrous collagens are aab or abc heterotrimers, yet few examples of heterotrimeric CMPs have been reported. Previously, CMP heterotrimers have only been accessible by total syntheses or by introducing complementary interstrand electrostatic or steric interactions. Here, we describe an abc CMP heterotrimer in which each contributing CMP consists of only three amino acids: glycine, proline and 4-hydroxyproline. Assembly of the heterotrimeric triple helix is directed by a combination of metal-ion coordination to set the relative register of the CMPs, and minimization of valence frustration to direct heterotrimerization. Assembly of the four-component mixture is facile and extremely rapid, and equilibration to the abc heterotrimer occurs within a few hours at modestly elevated temperatures. The melting temperatures of the metal-assembled collagen trimers are higher by some 30°C than the apopeptide assemblies. Two iterations of the design are described, and the outcomes suggest possibilities for designing self-assembling abc and abb heterotrimers.
| S-EPMC3202058 | biostudies-literature
Project description:The most abundant member of the collagen protein family, collagen I (also known as type I collagen; COL1), is composed of one unique (chain B) and two similar (chain A) polypeptides that self-assemble with one amino acid offset into a heterotrimeric triple helix. Given the offset, chain B can occupy either the leading (BAA), middle (ABA) or trailing (AAB) position of the triple helix, yielding three isomeric biomacromolecules with different protein recognition properties. Despite five decades of intensive research, there is no consensus on the position of chain B in COL1. Here, three triple-helical heterotrimers that each contain a putative von Willebrand factor (VWF) and discoidin domain receptor (DDR) recognition sequence from COL1 were designed with chain B permutated in all three positions. AAB demonstrated a strong preference for both VWF and DDR, and also induced higher levels of cellular DDR phosphorylation. Thus, we resolve this long-standing mystery and show that COL1 adopts an AAB register.
| S-EPMC7100791 | biostudies-literature
Project description:The de novo design of three protein chains that associate to form a heterotrimer (but not any of the possible two-chain heterodimers) and that can drive the assembly of higher-order branching structures is an important challenge for protein design. We designed helical heterotrimers with specificity conferred by buried hydrogen bond networks and large aromatic residues to enhance shape complementary packing. We obtained ten designs for which all three chains cooperatively assembled into heterotrimers with few or no other species present. Crystal structures of a helical bundle heterotrimer and extended versions, with helical repeat proteins fused to individual subunits, showed all three chains assembling in the designed orientation. We used these heterotrimers as building blocks to construct larger cyclic oligomers, which were structurally validated by electron microscopy. Our three-way junction designs provide new routes to complex protein nanostructures and enable the scaffolding of three distinct ligands for modulation of cell signaling.
| S-EPMC9758053 | biostudies-literature
Project description:Stability of the collagen triple helix is largely governed by its imino acid content, namely the occurrence of proline and 4R-hydroxyproline at the X and Y positions, respectively, of the periodic (Gly-X-Y)(n) sequence. Although other amino acids at these positions reduce stability of the triple helix, this can be partially compensated by introducing intermolecular side-chain salt bridges. This approach was previously used to design an abc-type heterotrimer composed of one basic, one acidic, and one neutral imino acid rich chain (Gauba and Hartgerink, J Am Chem Soc 2007;129:15034-15041). In this study, an abc-type heterotrimer was designed to be the most stable species using a sequence recombination strategy that preserved both the amino acid composition and the network of interchain salt bridges of the original design. The target heterotrimer had the highest T(m) of 50 °C, 7 °C greater than the next most stable species. Stability of the heterotrimer decreased with increasing ionic strength, consistent with the role of intermolecular salt bridges in promoting stability. Quantitative meta-analysis of these results and published stability measurements on closely related peptides was used to discriminate the contributions of backbone propensity and side-chain electrostatics to collagen stability.
| S-EPMC3557756 | biostudies-literature
Project description:Collagen model peptides are useful for understanding the assembly and structure of collagen triple helices. The design of self-assembling heterotrimeric helices is particularly challenging and often affords mixtures of non-covalent assemblies that are difficult to characterize by conventional NMR and CD spectroscopic techniques. This can render a detailed understanding of the factors that control heterotrimer formation difficult and restrict rational design. Here, we present a novel method based on electrospray ionization mass spectrometry to investigate homo- and heterotrimeric collagen model peptides. Under native conditions, the high resolving power of mass spectrometry was used to access the stoichiometric composition of different triple helices in complex mixtures. A temperature-controlled electrospray ionization source was built to perform thermal denaturation experiments and provided melting temperatures of triple helices. These were found to be in good agreement with values obtained from CD spectroscopic measurements. Importantly, for mixtures of coexisting homo- and heterotrimers, which are difficult to analyze by conventional methods, our technique allowed for the identification and monitoring of the unfolding of each individual species. Their respective melting temperatures could easily be accessed in a single experiment, using small amounts of sample.
| S-EPMC6977553 | biostudies-literature
Project description:RNA seq on primary pancreatic cancer (KPPC;Col1pdxKO cancer-collagen1 knockout) cells growing on collagen 1 homotrimers and heterotrimers. In this study, we identify that pancreatic cancer cells produce a unique Collagen1 homotrimer variant, in contrast to the Collagen1 heterotrimer produced by normal cells (such as fibroblasts). The variant forms of Collagen1 (homotrimer and heterotrimer) have distinct effects on cancer cell behaviors. We culture the pancreatic cancer cells on purified Collagen1 homotrimer and heterotrimer (as well as vehicle control) and examine the global changes of gene expression profile in cancer cells by variant Collagen1 subtypes.
2022-05-21 | GSE131666 | GEO
Project description:Hydroxyproline plays a major role in stabilizing collagenous domains in eukaryotic organisms. Lack of this modification is associated with significant lowering in the thermal stability of the collagen triple helix and may also affect fibrillogenesis and folding of the peptide chains. In contrast, even though bacterial collagens lack hydroxyproline, their thermal stability is comparable to that of fibrillar collagen. This has been attributed to the high frequency of charged amino acids found in bacterial collagen. Here we report a thermally stable hydroxyproline-free ABC heterotrimeric collagen mimetic system composed of decapositive and decanegative peptides and a zwitterionic peptide. None of the peptides contain hydroxyproline, and furthermore the zwitterionic peptide does not even contain proline. The heterotrimer is electrostatically stabilized via multiple interpeptide lysine-aspartate and lysine-glutamate salt bridges and maintains good thermal stability with a melting temperature of 37 °C. The ternary peptide mixture also populates a single composition ABC heterotrimer as confirmed by circular dichroism (CD) and nuclear magnetic resonance (NMR) spectroscopy. This system illustrates the power of axial salt bridges to direct and stabilize the self-assembly of a triple helix and may be useful in analogous designs in expression systems where the incorporation of hydroxyproline is challenging.
| S-EPMC3663077 | biostudies-literature
Project description:We have successfully designed an A:B:C collagen peptide heterotrimer using an automated computational approach. The algorithm maximizes the energy gap between the target and competing misfolded states while enforcing a minimum target stability. Circular dichroism (CD) measurements confirm that all three peptides are required to form a stable, structured triple helix. This study highlights the power of automated computational design, providing model systems to probe the biophysics of collagen assembly and developing general methods for the design of fibrous proteins.
| S-EPMC3443875 | biostudies-literature