Common Secondary Structure Motifs Research Articles

Mechanical unfolding of alpha- and beta-helical protein motifs.

Alpha-helices and beta-sheets are the two most common secondary structure motifs in proteins. Beta-helical structures merge features of the two motifs, containing two or three beta-sheet faces connected by loops or turns in a single protein. Beta-helical structures form the basis of proteins with diverse mechanical functions such as bacterial adhesins, phage cell-puncture devices, antifreeze proteins, and extracellular matrices. Alpha-helices are commonly found in cellular and extracellular matrix components, whereas beta-helices such as curli fibrils are more common as bacterial and biofilm matrix components. It is currently not known whether it may be advantageous to use one helical motif over the other for different structural and mechanical functions. To better understand the mechanical implications of using different helix motifs in networks, here we use Steered Molecular Dynamics (SMD) simulations to mechanically unfold multiple alpha- and beta-helical proteins at constant velocity at the single molecule scale. We focus on the energy dissipated during unfolding as a means of comparison between proteins and work normalized by protein characteristics (initial and final length, # H-bonds, # residues, etc.). We find that although alpha-helices such as keratin and beta-helices CsgA and CsgB can require similar amounts of work to unfold, the normalized work per hydrogen bond, initial end to end length, and number of residues is greater for beta-helices at the same pulling rate. To explain this, we analyze the orientation of the backbone alpha carbons and backbone hydrogen bonds during unfolding. We find that the larger width and shorter height of beta-helices results in smaller angles between the protein backbone and the pulling direction during unfolding. As subsequent strands are separated from the beta-helix core, the angle between the backbone and the pulling direction diminishes. This marks a transition where beta-sheet hydrogen bonds become loaded predominantly in a collective shearing mode, which requires a larger rupture force. This finding underlines the importance of geometry in optimizing resistance to mechanical unfolding in proteins. The helix radius is identified here as an important parameter that governs how much sacrificial energy dissipation capacity can be stored in protein networks, where beta-helices offer unique properties.

Examining the Stability of β-Sheets using the Charmm Drude Polarizable Force Field

β-Sheets are some of the most common secondary structure motifs in proteins, and are important for mediating protein-protein interactions through their association. This association can also lead to the aggregation of misfolded proteins into β-pleated-sheets in neurodegenerative disorders like Alzheimer's disease. Fixed charge, all-atom molecular dynamics simulations have adequately reproduced the folding free energy landscape of a small β-hairpin, the GB1 domain of protein G, which is a prototype for a larger β-pleated-sheet. Polarizable force fields should, in theory, be able to more accurately reproduce experimental results as they are a more realistic model of the molecular system. The CHARMM Drude model is a relatively efficient polarizable force field that has been shown to perform as well or better than traditional non-polarizable force fields in reproducing helical content and folding mechanisms of model α-helical peptides. To assess the model's accuracy for β-sheets, we examined the stability of the GB1 β-hairpin using the CHARMM Drude polarizable force field and two non-polarizable force fields, CHARMM36 and CHARMM22∗. Two-dimensional replica exchange umbrella sampling (REUS) simulations show that the β-hairpin is unstable in the Drude system, whereas it is either stable or quasistable in CHARMM36 and CHARMM22∗, respectively. The instability in Drude appears to be driven mostly by interactions between the peptide backbone and the surrounding water molecules. By tuning these interactions, we have shiftedthe stability back towards the β-hairpin state.

The Self‐Assembly of Helical Peptide Building Blocks

AbstractThe self‐assembly of peptide building blocks represents a central direction in bionanotechnology. A β‐sheet secondary structure serves as the basis of numerous peptide‐based bionanomaterials owing to their association with several neurodegenerative diseases. Although the α‐helix embodies the most common secondary structure motif in proteins, the utilization of this module as a building block for supramolecular aggregation is much less explored. In this Focus Review, we will discuss the advancement made so far to induce higher level organization in helical peptide building blocks. The primary design strategies involving coiled coil proteins mimetic approaches, incorporation of noncoded and synthetic amino acids and conjugation of peptides to aliphatic alkyl tails will be dealt with in detail. In parallel, we will also outline the self‐assembled materials developed in these progressions. Finally, possible future prospects of such assembly modules will be briefed.

Thermodynamics of β-Structures from Molecular Dynamics Simulations

β-Sheets are some of the most common secondary structure motifs in proteins, and are important for mediating protein-protein interactions through their association. This association can also lead to the aggregation of misfolded proteins into β-pleated-sheets in neurodegenerative disorders like amyloidosis. The folding pathway from random coil to β-sheet usually involves two competing process: (1) the collapse of a hydrophobic core, and (2) the formation of intrapeptide hydrogen bonds. It has been proposed, and shown computationally, that the hydrophobic core collapse precedes hydrogen bond formation. In this study we examine the thermodynamics of β-hairpin formation for the GB1 domain of protein G with molecular dynamics simulations by calculating a two-dimensional free energy surface in both vacuum and explicit water using as our reaction coordinates (1) the radius of gyration of the hydrophobic core and (2) the number of native hydrogen bonds, corresponding to the two aforementioned folding processes, respectively. We also compare the results of different versions of the CHARMM force field, namely CHARMM22, CHARMM22/CMAP, CHARMM22∗ and CHARMM36. Finally, we show how these methods can be applied to other β-structures in vivo, namely β-helix structures in the outer membrane of Gram-negative bacteria.

Packaging Signals in Two Single-Stranded RNA Viruses Imply a Conserved Assembly Mechanism and Geometry of the Packaged Genome

The current paradigm for assembly of single-stranded RNA viruses is based on a mechanism involving non-sequence-specific packaging of genomic RNA driven by electrostatic interactions. Recent experiments, however, provide compelling evidence for sequence specificity in this process both in vitro and in vivo. The existence of multiple RNA packaging signals (PSs) within viral genomes has been proposed, which facilitates assembly by binding coat proteins in such a way that they promote the protein–protein contacts needed to build the capsid. The binding energy from these interactions enables the confinement or compaction of the genomic RNAs. Identifying the nature of such PSs is crucial for a full understanding of assembly, which is an as yet untapped potential drug target for this important class of pathogens. Here, for two related bacterial viruses, we determine the sequences and locations of their PSs using Hamiltonian paths, a concept from graph theory, in combination with bioinformatics and structural studies. Their PSs have a common secondary structure motif but distinct consensus sequences and positions within the respective genomes. Despite these differences, the distributions of PSs in both viruses imply defined conformations for the packaged RNA genomes in contact with the protein shell in the capsid, consistent with a recent asymmetric structure determination of the MS2 virion. The PS distributions identified moreover imply a preferred, evolutionarily conserved assembly pathway with respect to the RNA sequence with potentially profound implications for other single-stranded RNA viruses known to have RNA PSs, including many animal and human pathogens.

Raman and Raman optical activity (ROA) analysis of RNA structural motifs

Raman and Raman optical activity (ROA) spectroscopies are well suited to the study of ribonucleic acids (RNA) as they are both sensitive to a large amount of structural information and applicable to most RNAs and experimental conditions. As such, they can fill the current gap between high- and low-resolution techniques currently used by RNA biologists. In this study we illustrate the complementarity of Raman and ROA spectroscopies by their application to two functional RNAs, a 37 nucleotide RNA fragment from Domain I of the encephalomyocarditis virus (EMCV) internal ribosome entry site (IRES) RNA and the L11 binding domain ribosomal RNA (LBD rRNA). Fingerprint spectra unique to the structure of each RNA are presented. We then discuss the Raman and ROA spectra of several oligonucleotides containing three common secondary structural motifs all found within the EMCV RNA; the GNRA tetraloop, pyrimidine-rich asymmetric bulge and a base mismatch. We show that highly detailed structural information can be obtained from these spectra.

Peptide β-Bend and 3 10-Helix: from 3D-Structural Studies to Applications as Templates

The 3 10-helix is a relatively common secondary structure motif in peptides and proteins. Its building block is one of various types of β-bend conformation which comprises an N α-acylated dipeptide alkylamide system. A complete 3D-structural characterization of this ternary helix has been achieved, thus allowing its unambiguous discrimination from the closely related α-helix. Recent applications of rigidified peptide β-bends and 3 10-helices as templates for investigations in synthetic organic chemistry (macrocyclization, catalysis), host–guest chemistry (molecular recognition), and physical chemistry (donor–acceptor interaction) will be discussed.

Binding of an aminoacridine derivative to a GAAA RNA tetraloop

RNA tetraloops are common secondary structural motifs in many RNAs, especially ribosomal RNAs. There are few studies of small molecule recognition of RNA tetraloops although tetraloops are known to interact with RNA receptors and proteins, and to form nucleation sites for RNA folding. In this paper, we investigate the binding of neomycin, kanamycin, 2,4-diaminoquinazoline, quinacrine, and an aminoacridine derivative (AD1) to a GAAA tetraloop using fluorescence spectroscopy. We have found that AD1 and quinacrine bind to the GAAA tetraloop with the highest affinity of the molecules examined. The equilibrium dissociation constant of the AD1–GAAA tetraloop complex was determined to be 1.6 μM. RNase I and lead acetate footprinting experiments suggested that AD1 binds to the junction between the loop and stem of the GAAA tetraloop.

Template-constrained cyclic peptide analogues of somatostatin: subtype-selective binding to somatostatin receptors and antiangiogenic activity

β-Turns are a common secondary structure motif found in proteins that play a role in protein folding and stability and participate in molecular recognition interactions. Somatostatin, a peptide hormone possessing a variety of therapeutically-interesting biological acitivities, contains a β-turn in its bioactive conformation. The β-turn and biological activities of somatostatin have been succesfully mimicked in cyclic hexapeptide analogues. Two novel, structured, non-peptidic molecules were developed that are capable of holding the bioactive tetrapeptide sequence of somatostatin analogues in a β-turn conformation, as measured by somatostatin receptor (SSTR) binding. Template-constrained cyclic peptides in which the ends of the -Tyr- d-Trp-Lys-Val-tetrapeptide were linked by scaffolds based on either an N,N′-dimethyl- N,N′-diphenylurea or a substituted biphenyl system ( DJS631 and DJS811, respectively), bound selectively to mouse SSTR2B and rat and human SSTR5 with affinities as high as 1 nM. DJS811, at a dose of 3 mg/kg/day, was shown in a mouse Matrigel model to inhibit angiogenesis to a level of 79%. The development of structured turn scaffolds allows β-turn sequences to be contained in the context of a compact structure, with less peptidic nature and potentially greater bioavailability than cyclic hexapeptides. These systems can be used to study the determinants of β-turn formation, as well as to probe the importance of turn sequences occurring in molecular recognition interactions. The antiangiogenic activity of DJS811 suggests that it may have antitumor activity as well. In addition, because SSTR2 is overexpressed on many types of tumors, DJS631 and DJS811 may be useful in the development of agents for tumor imaging or the radiotherapy of cancer.

Crystal structures of an A-form duplex with single-adenosine bulges and a conformational basis for site-specific RNA self-cleavage.

Bulged nucleotides are common secondary structural motifs in RNA molecules and are often involved in RNA-RNA and RNA-protein interactions. RNA is selectively cleaved at bulge sites (when compared to other sites within stems) in the presence of divalent metal cations. The effects of bulge nucleotides on duplex stability and topology have been extensively investigated, but no detailed X-ray structures of bulge-containing RNA fragments have been available. We have crystallized a self-complementary RNA-DNA chimeric 11-nucleotide sequence containing single-adenosine bulges under two different conditions, giving two distinct crystal forms. In both lattices the adenosines are looped out, leaving the stacking interactions in the duplex virtually unaffected. The bulges cause the duplex to kink in both cases. In one of the structures, the conformation of the bulged nucleotide places its modeled 2'-oxygen in line with the adjacent phosphate on the 3' side, where it is poised for nucleophilic attack. Single adenosine bulges cause a marked opening of the normally narrow RNA major groove in both crystal structures, rendering the bases more accessible to interacting molecules compared with an intact stem. The geometries around the looped-out adenosines are different in the two crystal forms, indicating that bulges can confer considerable local plasticity on the usually rigid RNA double helix. The results provide a conformational basis for the preferential, metal-assisted self-cleavage of RNA at bulged sites.