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Lifetime measurements reveal kinetic differences between homophilic cadherin bonds.

ABSTRACT: Cadherins are multidomain adhesion proteins whose interactions direct cell sorting during histogenesis. They determine cell adhesion specificity, but prior studies failed to identify physical differences that could underlie cell sorting. These single molecule studies identify kinetic and strength differences between different cadherins. They further demonstrate that the modular extracellular architecture of cleavage stage C-cadherin supports a multistate binding mechanism. These multiple bonds exhibit a kinetic hierarchy of strengths that map to the different cadherin domains. The outer two N-terminal domains of C-cadherin form two bound states with dissociation rates of 3.9 and 0.02 s(-1). The latter is 25-fold slower than between the corresponding epithelial cadherin segments. In addition to the two fast bonds, the five-domain fragment (CEC1-5) forms two additional stronger, longer-lived bonds with dissociation rates of 0.00039 and 0.00001 s(-1). We further quantified the lifetimes of bonds subject to a constant force, and thus identified multiple dissociation events with rates that agree quantitatively with the force spectroscopy data. The qualitative features are similar to those reported for epithelial cadherin. However, the significant differences in the dissociation rates of the outer domains, which include the specificity-determining region, suggest that kinetic differences may determine cadherin discrimination, rather than adhesion energies.

SUBMITTER: Bayas MV

PROVIDER: S-EPMC1367289 | biostudies-literature | 2006 Feb

REPOSITORIES: biostudies-literature

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Lifetime measurements reveal kinetic differences between homophilic cadherin bonds.

Bayas Marco V MV Leung Andrew A Evans Evan E Leckband Deborah D

Biophysical journal 20051202 4

Cadherins are multidomain adhesion proteins whose interactions direct cell sorting during histogenesis. They determine cell adhesion specificity, but prior studies failed to identify physical differences that could underlie cell sorting. These single molecule studies identify kinetic and strength differences between different cadherins. They further demonstrate that the modular extracellular architecture of cleavage stage C-cadherin supports a multistate binding mechanism. These multiple bonds e ...[more]

PMID: 16326909

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Project description:Sphingomyelin is an abundant lipid in some cellular membrane domains, such as lipid rafts. Hydrogen bonding and hydrophobic interactions of the lipid with surrounding components such as neighboring sphingomyelin and cholesterol (Cho) are widely considered to stabilize the raft-like liquid-ordered (Lo) domains in membrane bilayers. However, details of their interactions responsible for the formation of Lo domains remain largely unknown. In this study, the enantiomer of stearoyl sphingomyelin (ent-SSM) was prepared, and its physicochemical properties were compared with the natural SSM and the diastereomer of SSM to examine possible stereoselective lipid-lipid interactions. Interestingly, differential scanning calorimetry experiments demonstrated that palmitoyl sphingomyelin, with natural stereochemistry, exhibited higher miscibility with SSM bilayers than with ent-SSM bilayers, indicating that the homophilic sphingomyelin interactions occurred in a stereoselective manner. Solid-state 2H NMR revealed that Cho elicited its ordering effect very similarly on SSM and ent-SSM (and even on the diastereomer of SSM), suggesting that SSM-Cho interactions are not significantly affected by stereospecific hydrogen bonding. SSM and ent-SSM formed gel-like domains with very similar lateral packing in SSM/Cho/palmitoyloleoyl phosphatidylcholine membranes, as shown by fluorescence lifetime experiments. This observation can be explained by a homophilic hydrogen-bond network, which was largely responsible for the formation of gel-like nanodomains of SSMs (or ent-SSM). Our previous study revealed that Cho-poor gel-like domains contributed significantly to the formation of an Lo phase in sphingomyelin/Cho membranes. The results of the study presented here further show that SSM-SSM interactions occur near the headgroup region, whereas hydrophobic SSM-Cho interactions appeared important in the bilayer interior for Lo domain formation. The homophilic interactions of sphingomyelins could be mainly responsible for the formation of the domains of nanometer size, which may correspond to the small sphingomyelin/Cho-based rafts that temporally occur in biological membranes.

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Lifetime measurements reveal kinetic differences between homophilic cadherin bonds.

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Lifetime measurements reveal kinetic differences between homophilic cadherin bonds.

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