Identification of hydrophobic interactions between relaxin-3 and its receptor RXFP3: implication for a conformational change in the B-chain C-terminus during receptor binding

CD44 is the receptor for hyaluronan (HA) and mediates cell rolling under fluid shear stress. The HA-binding domain (HABD) of CD44 interconverts between a low-affinity, ordered (O) state and a high-affinity, partially disordered (PD) state, by the conformational change of the C-terminal region, which is connected to the plasma membrane. To examine the role of tensile force on CD44-mediated rolling, we used a cell-free rolling system, in which recombinant HABDs were attached to beads through a C-terminal or N-terminal tag. We found that the rolling behavior was stabilized only at high shear stress, when the HABD was attached through the C-terminal tag. In contrast, no difference was observed for the beads coated with HABD mutants that constitutively adopt either the O state or the PD state. Steered molecular dynamics simulations suggested that the force from the C terminus disrupts the interaction between the C-terminal region and the core of the domain, thus providing structural insights into how the mechanical force triggers the allosteric O-to-PD transition. Based on these results, we propose that the force applied from the C terminus enhances the HABD–HA interactions by inducing the conformational change to the high-affinity PD transition more rapidly, thereby enabling CD44 to mediate lymphocyte trafficking and hematopoietic progenitor cell homing under high-shear conditions.

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Limited and Defined Truncation at the C Terminus Enhances Receptor Binding and Degranulation Activity of the Neutrophil-activating Peptide 2 (NAP-2)

Journal of Biological Chemistry ◽

10.1074/jbc.270.11.6338 ◽

1995 ◽

Vol 270 (11) ◽

pp. 6338-6344 ◽

Cited By ~ 17

Author(s):

Jan E. Ehlert ◽

Frank Petersen ◽

Michael H. G. Kubbutat ◽

Johannes Gerdes ◽

Hans-Dieter Flad ◽

...

Keyword(s):

Receptor Binding ◽

C Terminus

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Degradation of ornithine decarboxylase: exposure of the C-terminal target by a polyamine-inducible inhibitory protein

Molecular and Cellular Biology ◽

10.1128/mcb.13.4.2377-2383.1993 ◽

1993 ◽

Vol 13 (4) ◽

pp. 2377-2383

Author(s):

X Li ◽

P Coffino

Keyword(s):

Conformational Change ◽

Ornithine Decarboxylase ◽

Binding Protein ◽

Active Form ◽

Feedback Regulation ◽

Degradation Process ◽

Protein Inhibitor ◽

Inhibitory Protein ◽

C Terminus ◽

N Terminus

Polyamine-mediated degradation of vertebrate ornithine decarboxylase (ODC) is associated with the production of antizyme, a reversible tightly binding protein inhibitor of ODC activity. The interaction of antizyme with a binding element near the N terminus of ODC is essential but not sufficient for regulation of the enzyme by polyamines (X. Li and P. Coffino, Mol. Cell. Biol. 12:3556-2562, 1992). We now show that a second element present at the C terminus is required for the degradation process. Antizyme caused a conformational change in ODC, which made the C terminus of ODC more accessible. Blocking the C terminus with antibody prevented degradation. Tethering the C terminus by creating a circularly permuted, enzymatically active form of ODC prevented antizyme-mediated degradation. These data elucidate a form of feedback regulation whereby excess polyamines induce destruction of ODC, the enzyme that initiates their biosynthesis.

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Residue-Level Contact Reveals Modular Domain Interactions of PICK1 Are Driven by Both Electrostatic and Hydrophobic Forces

Frontiers in Molecular Biosciences ◽

10.3389/fmolb.2020.616135 ◽

2021 ◽

Vol 7 ◽

Author(s):

Amy O. Stevens ◽

Yi He

Keyword(s):

Hydrophobic Interactions ◽

Pdz Domain ◽

Md Simulations ◽

Intramolecular Interactions ◽

Coarse Grained ◽

Scaffolding Protein ◽

Concave Surface ◽

Stable Complex ◽

Bar Domain ◽

C Terminus

PICK1 is a multi-domain scaffolding protein that is uniquely comprised of both a PDZ domain and a BAR domain. While previous experiments have shown that the PDZ domain and the linker positively regulate the BAR domain and the C-terminus negatively regulates the BAR domain, the details of internal regulation mechanisms are unknown. Molecular dynamics (MD) simulations have been proven to be a useful tool in revealing the intramolecular interactions at atomic-level resolution. PICK1 performs its biological functions in a dimeric form which is extremely computationally demanding to simulate with an all-atom force field. Here, we use coarse-grained MD simulations to expose the key residues and driving forces in the internal regulations of PICK1. While the PDZ and BAR domains do not form a stable complex, our simulations show the PDZ domain preferentially interacting with the concave surface of the BAR domain over other BAR domain regions. Furthermore, our simulations show that the short helix in the linker region can form interactions with the PDZ domain. Our results reveal that the surface of the βB-βC loop, βC strand, and αA-βD loop of the PDZ domain can form a group of hydrophobic interactions surrounding the linker helix. These interactions are driven by hydrophobic forces. In contrast, our simulations reveal a very dynamic C-terminus that most often resides on the convex surface of the BAR domain rather than the previously suspected concave surface. These interactions are driven by a combination of electrostatic and hydrophobic interactions.

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Structural and functional aspects of the multiplicity of Neu differentiation factors

Molecular and Cellular Biology ◽

10.1128/mcb.14.3.1909-1919.1994 ◽

1994 ◽

Vol 14 (3) ◽

pp. 1909-1919

Author(s):

D Wen ◽

S V Suggs ◽

D Karunagaran ◽

N Liu ◽

R L Cupples ◽

...

Keyword(s):

Receptor Binding ◽

Transmembrane Domain ◽

Structural Heterogeneity ◽

Binding Activity ◽

Biologically Active ◽

Specific Rate ◽

C Terminus ◽

Functional Aspects ◽

Differentiation Factors ◽

Variable Domains

We used molecular cloning and functional analyses to extend the family of Neu differentiation factors (NDFs) and to explore the biochemical activity of different NDF isoforms. Exhaustive cloning revealed the existence of six distinct fibroblastic pro-NDFs, whose basic transmembrane structure includes an immunoglobulin-like motif and an epidermal growth factor (EGF)-like domain. Structural variation is confined to three domains: the C-terminal portion of the EGF-like domain (isoforms alpha and beta), the adjacent juxtamembrane stretch (isoforms 1 to 4), and the variable-length cytoplasmic domain (isoforms a, b, and c). Only certain combinations of the variable domains exist, and they display partial tissue specificity in their expression: pro-NDF-alpha 2 is the predominant form in mesenchymal cells, whereas pro-NDF-beta 1 is the major neuronal isoform. Only the transmembrane isoforms were glycosylated and secreted as biologically active 44-kDa glycoproteins, implying that the transmembrane domain functions as an internal signal peptide. Extensive glycosylation precedes proteolytic cleavage of pro-NDF but has no effect on receptor binding. By contrast, the EGF-like domain fully retains receptor binding activity when expressed separately, but its beta-type C terminus displays higher affinity than alpha-type NDFs. Likewise, structural heterogeneity of the cytoplasmic tails may determine isoform-specific rate of pro-NDF processing. Taken together, these results suggest that different NDF isoforms are generated by alternative splicing and perform distinct tissue-specific functions.

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Structural and Biochemical Characterization of Botulinum Neurotoxin Subtype B2 Binding to Its Receptors

Toxins ◽

10.3390/toxins12090603 ◽

2020 ◽

Vol 12 (9) ◽

pp. 603

Author(s):

Jonathan R. Davies ◽

Geoffrey Masuyer ◽

Pål Stenmark

Keyword(s):

Receptor Binding ◽

Molecular Mechanisms ◽

Biochemical Characterization ◽

Hydrophobic Interactions ◽

Binding Assays ◽

Protein Receptor ◽

Wide Range ◽

Protein Receptors ◽

Therapeutic Molecules ◽

Botulinum Neurotoxins

Botulinum neurotoxins (BoNTs) can be used therapeutically to treat a wide range of neuromuscular and neurological conditions. A collection of natural BoNT variants exists which can be classified into serologically distinct serotypes (BoNT/B), and further divided into subtypes (BoNT/B1, B2, …). BoNT subtypes share a high degree of sequence identity within the same serotype yet can display large variation in toxicity. One such example is BoNT/B2, which was isolated from Clostridium botulinum strain 111 in a clinical case of botulism, and presents a 10-fold lower toxicity than BoNT/B1. In an effort to understand the molecular mechanisms behind this difference in potency, we here present the crystal structures of BoNT/B2 in complex with the ganglioside receptor GD1a, and with the human synaptotagmin I protein receptor. We show, using receptor-binding assays, that BoNT/B2 has a slightly higher affinity for GD1a than BoNT/B1, and confirm its considerably weaker affinity for its protein receptors. Although the overall receptor-binding mechanism is conserved for both receptors, structural analysis suggests the lower affinity of BoNT/B2 is the result of key substitutions, where hydrophobic interactions important for synaptotagmin-binding are replaced by polar residues. This study provides a template to drive the development of future BoNT therapeutic molecules centered on assessing the natural subtype variations in receptor-binding that appears to be one of the principal stages driving toxicity.

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A change-in-hand mechanism for S100 signalling

Biochemistry and Cell Biology ◽

10.1139/o98-062 ◽

1998 ◽

Vol 76 (2-3) ◽

pp. 324-333 ◽

Cited By ~ 36

Author(s):

Steven P Smith ◽

Gary S Shaw

Keyword(s):

Conformational Change ◽

Calcium Ion ◽

Calcium Binding ◽

Binding Proteins ◽

Three Dimensional ◽

Calcium Binding Proteins ◽

C Terminus ◽

Ef Hand ◽

Sensor Proteins ◽

Binding Loop

S100 proteins are a group of small dimeric calcium-binding proteins making up a large subclass of the EF-hand family of calcium-binding proteins. Members of this family of proteins have been proposed to act as intracellular calcium modulatory proteins in a fashion analogous to that of the EF-hand sensor proteins troponin-C and calmodulin. Recently, NMR spectroscopy has provided the three-dimensional structures of the S100 family members S100A6 and S100B in both the apo- and calcium-bound forms. These structures have allowed for the identification of a novel calcium-induced conformational change termed the change-in-hand mechanism. Helix III of the C-terminal calcium-binding loop changes its helix-helix interactions (or handness) with the remainder of the molecule primarily owing to the reorientation of the backbone in an effort to coordinate the calcium ion. This reorientation of helix III exposes several residues in the C-terminus and linker regions of S100B resulting in the formation of a hydrophobic patch surrounded be a number of acidic residues. This site is the proposed region for protein-protein recognition.Key words: S100, calcium-binding protein, EF-hand, conformational change.

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Structural and Functional Features of the Reovirus σ1 Tail

Journal of Virology ◽

10.1128/jvi.00336-18 ◽

2018 ◽

Vol 92 (14) ◽

Cited By ~ 15

Author(s):

Melanie H. Dietrich ◽

Kristen M. Ogden ◽

Jacob M. Long ◽

Rebecca Ebenhoch ◽

Alexandra Thor ◽

...

Keyword(s):

Conformational Change ◽

Receptor Binding ◽

Coiled Coil ◽

Chloride Ions ◽

The Body ◽

Full Length ◽

Cell Entry ◽

Target Cells ◽

Attachment Protein ◽

Functional Features

ABSTRACT Mammalian orthoreovirus attachment to target cells is mediated by the outer capsid protein σ1, which projects from the virion surface. The σ1 protein is a homotrimer consisting of a filamentous tail, which is partly inserted into the virion; a body domain constructed from β-spiral repeats; and a globular head with receptor-binding properties. The σ1 tail is predicted to form an α-helical coiled coil. Although σ1 undergoes a conformational change during cell entry, the nature of this change and its contributions to viral replication are unknown. Electron micrographs of σ1 molecules released from virions identified three regions of flexibility, including one at the midpoint of the molecule, that may be involved in its structural rearrangement. To enable a detailed understanding of essential σ1 tail organization and properties, we determined high-resolution structures of the reovirus type 1 Lang (T1L) and type 3 Dearing (T3D) σ1 tail domains. Both molecules feature extended α-helical coiled coils, with T1L σ1 harboring central chloride ions. Each molecule displays a discontinuity (stutter) within the coiled coil and an unexpectedly seamless transition to the body domain. The transition region features conserved interdomain interactions and appears rigid rather than highly flexible. Functional analyses of reoviruses containing engineered σ1 mutations suggest that conserved residues predicted to stabilize the coiled-coil-to-body junction are essential for σ1 folding and encapsidation, whereas central chloride ion coordination and the stutter are dispensable for efficient replication. Together, these findings enable modeling of full-length reovirus σ1 and provide insight into the stabilization of a multidomain virus attachment protein. IMPORTANCE While it is established that different conformational states of attachment proteins of enveloped viruses mediate receptor binding and membrane fusion, less is understood about how such proteins mediate attachment and entry of nonenveloped viruses. The filamentous reovirus attachment protein σ1 binds cellular receptors; contains regions of predicted flexibility, including one at the fiber midpoint; and undergoes a conformational change during cell entry. Neither the nature of the structural change nor its contribution to viral infection is understood. We determined crystal structures of large σ1 fragments for two different reovirus serotypes. We observed an unexpectedly tight transition between two domains spanning the fiber midpoint, which allows for little flexibility. Studies of reoviruses with engineered changes near the σ1 midpoint suggest that the stabilization of this region is critical for function. Together with a previously determined structure, we now have a complete model of the full-length, elongated reovirus σ1 attachment protein.

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