Open Hinge Research Articles

The Barrette

The Barrette Carling McManus (bio) She couldn't have known—her fingers brushing my forehead,pulling loose strands from my faceto fasten a plastic clasp, a butterfly,pressed hard against my skulluntil the loud snap sounded insideher bedroom where she eyed our familybefore church. "That's better," she'd sayto the mirror, palming my shoulders.We'd stand as if for a family photo,a mother with her eldest daughter,the bright barrette biting my scalp.No way she would have knownthat these insects scavenge saltfrom rancid animal skin and with rolled,forked tongues, suck nectar from fatflower heads, lick the acidic surfacesof polluted pools, flit in the air untillanding on a lizard's eyelid to drinkits tears. Of course she didn't know—not even the British book binderwho hand-stitched the first volumeof Butterflies and Moths understoodexactly how much nature affixeselegance to deviance, like a pairof wings folded in an open hinge. [End Page 13] Carling McManus Carling McManus is a queer poet whose work appears or is forthcoming in Meridian, Best New Poets, The Beloit Poetry Journal, and was selected as the Poetry Prizewinner in Carve Magazine's 2021 Prose & Poetry Contest. She is a two-time Rona Jaffe Foundation Scholar for the Bread Loaf Writers' Conference and a Community of Writers alum. Carling lives on a mountainside with her wife and two border collies in Mink Shoals, West Virginia. Copyright © 2022-2023 Pleiades and Pleiades Press

Colicin E1 opens its hinge to plug TolC.

The double membrane architecture of Gram-negative bacteria forms a barrier that is impermeable to most extracellular threats. Bacteriocin proteins evolved to exploit the accessible, surface-exposed proteins embedded in the outer membrane to deliver cytotoxic cargo. Colicin E1 is a bacteriocin produced by, and lethal to, Escherichia coli that hijacks the outer membrane proteins (OMPs) TolC and BtuB to enter the cell. Here, we capture the colicin E1 translocation domain inside its membrane receptor, TolC, by high-resolution cryo-electron microscopy to obtain the first reported structure of a bacteriocin bound to TolC. Colicin E1 binds stably to TolC as an open hinge through the TolC pore-an architectural rearrangement from colicin E1's unbound conformation. This binding is stable in live E. coli cells as indicated by single-molecule fluorescence microscopy. Finally, colicin E1 fragments binding to TolC plug the channel, inhibiting its native efflux function as an antibiotic efflux pump, and heightening susceptibility to three antibiotic classes. In addition to demonstrating that these protein fragments are useful starting points for developing novel antibiotic potentiators, this method could be expanded to other colicins to inhibit other OMP functions.

Colicin E1 binds to TolC as an open hinge to penetrate the outer-membrane

The double membrane architecture of Gram-negative bacteria forms a barrier that is effectively impermeable to extracellular threats. Bacteriocin proteins evolved to exploit the accessible, surface-exposed proteins embedded in the outer membrane to deliver cytotoxic cargo. Colicin E1 is a bacteriocin produced by, and lethal to, Escherichia coli that hijacks the outer membrane proteins TolC and BtuB to enter the cell. Here we capture the colicin E1 translocation domain inside its membrane receptor, TolC, by high-resolution cryoEM, the first reported structure of a bacteriocin bound to TolC.

Floquet chiral hinge modes and their interplay with Weyl physics in a three-dimensional lattice

We demonstrate that a three dimensional time-periodically driven (Floquet) lattice can exhibit chiral hinge states and describe their interplay with Weyl physics. A peculiar type of the hinge states are enforced by the repeated boundary reflections with lateral Goos-H\"{a}nchen like shifts occurring at the second-order boundaries of our system. Such chiral hinge modes coexist in a wide range of parameters regimes with Fermi arc surface states connecting a pair of Weyl points in a two-band model. We find numerically that these modes still preserve their locality along the hinge and their chiral nature in the presence of local defects and other parameter changes. We trace the robustness of such chiral hinge modes to special band structure unique in a Floquet system allowing all the eigenstates to be localized in quasi-one-dimensional regions parallel to each other when open hinge boundaries are introduced. The implementation of a model featuring both the second-order Floquet skin effect and the Weyl physics is straightforward with ultracold atoms in optical superlattices.

A novel class of selective CK2 inhibitors targeting its open hinge conformation

Protein kinase CK2 sustains cancer growth, especially in hematological malignancies. Its inhibitor SRPIN803, based on a 6-methylene-5-imino-1,3,4-thiadiazolopyrimidin-7-one scaffold, showed notable specificity. Our synthesis of the initially proposed SRPIN803 resulted in its constitutional isomer SRPIN803-revised, where the 2-cyano-2-propenamide group does not cyclise and fuse to the thiadiazole ring. Its crystallographic structure in complex with CK2α identifies the structural determinants of the reported specificity. SRPIN803-revised explores the CK2 open hinge conformation, extremely rare among kinases, also interacting with side chains from this region. Its optimization lead to the more potent compound 4, which inhibits endocellular CK2, significantly affects viability of tumour cells and shows remarkable selectivity on a panel of 320 kinases.

Antibody-mediated immune exclusion of HIV.

Although approximately 90% of all HIV transmissions in humans occur through mucosal contact, the induction of mucosal anti-HIV immune responses has remained understudied. Here we summarize data demonstrating the powerful protection that is achievable at mucosal frontlines through virus-specific mucosal IgA alone or combined with IgG. Passive immunization with different monoclonal antibody subclasses but identical epitope specificity (the conserved V3-loop crown of HIV gp120) has revealed that the dimeric IgA1 (dIgA1) form with its open hinge can prevent simian-human immunodeficiency virus (SHIV) acquisition in rhesus macaques at a higher rate than dIgA2. Both dIgAs neutralized the challenge SHIV equally well. Protection was linked to better virion capture and inhibition of cell-free virus transcytosis by dIgA1. Synergistic interactions at the mucosal level between the IgG1 and dIgA2 versions of this monoclonal antibody yielded complete protection. Active vaccine strategies designed to induce mucosal IgA and systemic/mucosal IgG have given promising data. This review seeks to highlight the importance of mucosal IgAs in preventing virus acquisition. Passive immunization gave proof-of-concept for immune exclusion by mucosally administered monoclonal dIgAs. Unanswered questions remain regarding the interplay between mucosal IgA and other host immune defenses, including their induction with active immunization.

Mechanism of flexibility control for ATP access of hepatitis C virus NS3 helicase

Hepatitis C virus (HCV) NS3 helicase couples adenosine triphosphate (ATP) binding and hydrolysis to polynucleotide unwinding. Understanding the regulation mechanism of ATP binding will facilitate targeting of the ATP-binding site for potential therapeutic development for hepatitis C. T324, an amino acid residue connecting domains 1 and 2 of NS3 helicase, has been suggested as part of a flexible hinge for opening of the ATP-binding cleft, although the detailed mechanism remains largely unclear. We used computational simulation to examine the mutational effect of T324 on the dynamics of the ATP-binding site. A mutant model, T324A, of the NS3 helicase apo structure was created and energy was minimized. Molecular dynamics simulation was conducted for both wild type and the T324A mutant apo structures to compare their differences. For the mutant structure, histogram analysis of pairwise distances between residues in domains 1 and 2 (E291-Q460, K210-R464 and R467-T212) showed that separation between the two domains was reduced by ∼10% and the standard deviation by ∼33%. Root mean square fluctuation (RMSF) analysis demonstrated that residues in close proximity to residue 324 have at least 30% RMSF value reductions in the mutant structure. Solvent RMSF analysis showed that more water molecules were trapped near D290 and H293 in domain 1 to form an extensive interaction network constraining cleft opening. We also demonstrated that the T324A mutation established a new atomic interaction with V331, revealing that an atomic interaction cascade from T324 to residues in domains 1 and 2 controls the flexibility of the ATP-binding cleft.

Homology models for domains 21–23 of human tropoelastin shed light on lysine crosslinking

The contiguous crosslinking domain at the center of human tropoelastin encoded by exons 21–23 contains an unusual ‘hinge’ region thought to adopt both open and closed conformations. Key lysines 425 and 437 have been implicated in both artificial and lysyl oxidase mediated crosslinks. We have examined the importance of hinge conformation to the proximity of these lysines and their ability to undergo intramolecular and intermolecular crosslinks using homology models. The results, counter intuitively, indicate that the more open hinge conformations favor intramolecular crosslinking, while the more closed conformations favor intermolecular crosslinking. We also present evidence that the sidechains of lysines 425 and 437 are able to make direct contact enabling an intramolecular lysyl oxidase mediated crosslink.

There's a catch

Leukocytes are more likely to grab onto a vascular surface when under flow than in the absence of flow. This counterintuitive behavior relies on bonds between transmembrane selectins on the blood cell and their vascular ligands. These so-called “catch bonds” get stronger because flow flips open selectin into a binding-ready conformation, Lou et al. show on page 1107. Figure 1 Low force (left) favors dissociation of selectin (pink) from its ligand (blue). More force opens the selectin hinge and allows the ligand to slide into new interactions (right). The authors have solved the structure of L-selectin, which can now be added to previous structures of P- and E-selectin. The extracellular domains of all three have an N-terminal lectin domain and an EGF-like domain that are separated by a hinge. In L- and E-selectin, the two sides of this hinge are connected by a hydrogen bond between a tyrosine at residue 37 and an asparagine at residue 138. In the absence of force, the lectin domain of L-selectin flipped between closed and open angles relative to the EGF domain, flexing at the hinge. But with increasing force, L-selectin was more often in an open conformation. A mutant L-selectin without the hydrogen bond between Y37 and N138 took less than half the force to be opened than did the wild type, suggesting that the hinge is held shut by this hydrogen bond. Maximal tethering of microspheres with this mutant L-selectin also occurred at a lower flow. The change was due to increased rotational freedom of the mutant's hinge, which increased the likelihood of protein–ligand contact. After the L-selectin bound its ligand, the authors found, dissociation also slowed as force increased (up to an optimal level, at which point the selectins are fully open). Again the optimal force was lower for the hydrogen bond mutant than for wild-type L-selectin. The group proposes that an open hinge, which aligns selectin's binding interface with the direction of flow, might allow a ligand that would otherwise let go to slide along its selectin until it can rebind to a new site. P-selectin has a glycine at position 138 and thus does not form a hinge-closing hydrogen bond. The increased flexibility of the P-selectin hinge is consistent with its ability to form catch bonds under smaller shear forces than those of L-selectin.

Domain swapping in N-truncated human cystatin C.

Human cystatin C (HCC) inhibits papain-like cysteine proteases by a binding epitope composed of two beta-hairpin loops and the N-terminal segment. HCC is found in all body fluids and is present at a particularly high level in the cerebrospinal fluid. Oligomerization of HCC leads to amyloid deposits in brain arteries at advanced age but this pathological process is greatly accelerated with a naturally occurring Leu68Gln variant, resulting in fatal amyloidosis in early adult life. When proteins are extracted from human cystatin C amyloid deposits, an N-terminally truncated cystatin C (THCC) is found, lacking the first ten amino acid residues of the native sequence. It has been shown that the cerebrospinal fluid may cause this N-terminal truncation, possibly because of disintegration of the leucocytes normally present in this fluid, and the release of leucocyte proteolytic enzymes. HCC is the first disease-causing amyloidogenic protein for which oligomerization via 3D domain swapping has been observed. The aggregates arise in the crystallization buffer and have the form of 2-fold symmetric dimers in which a long alpha-helix of one molecule, flanked by two adjacent beta-strands, has replaced an identical domain of the other molecule, and vice versa. Consistent with a conformational change at one of the beta-hairpin loops of the binding epitope, the dimers (and also any other oligomers, including amyloid aggregates) are inactive as papain inhibitors. Here, we report the structure of N-truncated HCC, the dominant form of cystatin C in amyloid deposits. Although the protein crystallized under conditions that are drastically different from those for the full-length protein, the structure reveals dimerization by the same act of domain swapping. However, the new crystal structure is composed of four independent HCC dimers, none of which has the exact 2-fold symmetry of the full-length dimer. While the four dimers have the same overall topology, the exact relation between the individual domains shows a variability that reflects the flexibility at the dimer-specific open interface, which in the case of 3D domain-swapped HCC consists of beta-interactions between the open hinge loops and results in an unusually long intermolecular beta-sheet. The dimers are engaged in further quaternary interactions resulting in spherical, closed octameric assemblies that are identical to that present in the crystal of the full-length protein. The octamers interact via hydrophobic patches formed on the surface of the domain-swapped dimers as well as by extending the dimer beta-sheet through intermolecular contacts.

Molecular dynamics simulations of DNA curvature and flexibility: helix phasing and premelting.

Recent studies of DNA axis curvature and flexibility based on molecular dynamics (MD) simulations on DNA are reviewed. The MD simulations are on DNA sequences up to 25 base pairs in length, including explicit consideration of counterions and waters in the computational model. MD studies are described for ApA steps, A-tracts, for sequences of A-tracts with helix phasing. In MD modeling, ApA steps and A-tracts in aqueous solution are essentially straight, relatively rigid, and exhibit the characteristic features associated with the B'-form of DNA. The results of MD modeling of A-tract oligonucleotides are validated by close accord with corresponding crystal structure results and nuclear magnetic resonance (NMR) nuclear Overhauser effect (NOE) and residual dipolar coupling (RDC) structures of d(CGCGAATTCGCG) and d(GGCAAAAAACGG). MD simulation successfully accounts for enhanced axis curvature in a set of three sequences with phased A-tracts studied to date. The primary origin of the axis curvature in the MD model is found at those pyrimidine/purine YpR "flexible hinge points" in a high roll, open hinge conformational substate. In the MD model of axis curvature in a DNA sequence with both phased A-tracts and YpR steps, the A-tracts appear to act as positioning elements that make the helix phasing more precise, and key YpR steps in the open hinge state serve as curvature elements. Our simulations on a phased A-tract sequence as a function of temperature show that the MD simulations exhibit a premelting transition in close accord with experiment, and predict that the mechanism involves a B'-to-B transition within A-tracts coupled with the prediction of a transition in key YpR steps from the high roll, open hinge, to a low roll, closed hinge substate. Diverse experimental observations on DNA curvature phenomena are examined in light of the MD model with no serious discrepancies. The collected MD results provide independent support for the "non-A-tract model" of DNA curvature. The "junction model" is indicated to be a special case of the non-A-tract model when there is a Y base at the 5' end of an A-tract. In accord with crystallography, the "ApA wedge model" is not supported by MD.

Quantitative sampler for collecting invertebrates associated with submersed and floating-leaved macrophytes

A new hand-operated sampler was developed consisting of a perspex cylinder (thickness 0.5 cm, length 32 cm, diameter 13 cm) cut in half lengthwise. The valves are joined together by means of a piano hinge for opening and closing of the sampler. In each of the sites there are openings covered with a net of 0.18×0.18 mm mesh. The external free edges of the halves are covered with weather stripping. The apparatus allows sampling of epiphytic fauna, or animals swimming around various macrophyte structures, including those trailing on the bottom, and those with floating leaves.

Reasoning about the referent of a picture versus reasoning about the picture as the referent: an effect of visual realism.

Research on picture perception and picture-based problem solving has generally considered the information that enables one to "see" and think about a picture's subject matter. However, people often reason about a picture or representation as the referent itself. The question addressed here is whether pictorial features themselves help determine when one reasons about the referent of an image, as with an engrossing movie, and when one reasons about the image in its own right, as with abstract art. Two experiments tested the hypothesis that pictures with relatively high fidelity to their referents lead people to think about those referents, whereas pictures with relatively low fidelity lead people to think about the picture as a referent. Subjects determined whether marks on the bottom and top boards of an open hinge would meet if the hinge were closed. Accuracy and latency results indicated that subjects who saw realistic displays simulated the physical behavior of the hinge through analog imagery. In contrast, subjects who saw schematic displays tended to reason about static features of the display such as line lengths and angles. The results demonstrate that researchers must be cautious when generalizing from reasoning about diagrammatic materials to reasoning about the referents themselves.

Alteration of the conformation of human IgG subclasses by reduction of the hinge SS bonds

Human IgG changed molecular size upon mild reduction and alkylation as shown by HPLC gel filtration. IgG1, IgG2 and IgG4 proteins increased in molecular size while IgG3 proteins were decreased in molecular size by this treatment. Several proteins within each subclass covering different light chain types and Gm types were tested all showing the same effect. A plausible explanation was related to the hinge and to the C H2 region since Fab fragments experienced unchanged molecular size irrespective of IgG subclass while Fc (of IgG1, IgG2, IgG3, containing only two aa of the 62 aa long hinge of IgG3 and IgG4) increased in size and Fch (which contains most of the 62 aa long hinge region of IgG3) decreased in size upon reduction and alkylation. It is postulated that reduction of the hinge SS bonds permit the IgG and Fc molecules to open up in the c h 2 region due to the lack of trans-interaction here, resulting in a larger molecular size. For IgG3 and Fch (from IgG3) molecules there was an opposite and even greater effect on the open polyproline like structure of the γ 3 hinge which depends on intact SS bonds (there are 11 bonds here). Reduction of these SS bonds apparently breaks down this open hinge structure resulting in a net decrease in molecular size of IgG3 and Fch molecules.