Intertwined αβ Spectrin Meeting Helical Actin Prot

Authors: Paul Sche Carlos Vera L Amy Sung

Publish Date: 2011/03/17

Volume: 39, Issue: 7, Pages: 1984-

PDF Link

Abstract

Our 3D model for a junctional complex JC in the erythrocyte membrane skeleton proposed that the helical actin protofilament functions as a mechanical axis for three pairs of αβ spectrin Sp and each pair wraps around the protofilament in a backtoback fashion The distal end of each Sp is further associated with the lipid bilayer by a suspension complex SC Here we detail how splitting and rejoining of αβ Sp around a protofilament may form a loop that sustains and equilibrates tension Sequential association of β and α Sp solves the challenge of constructing multiple loops along the protofilament and topological connection facilitates their reassociation The wraparound model minimizes the strain of the actin binding site on β Sp due to tension redirection or sliding of intertwined Sp Pairing Sp balances the opposing forces and provides a mechanism for elastic recovery The wraparound junction thus provides mechanical advantages over a pointattachment junction in maintaining the integrity and functionality of the network Severing α or β Sp may convert a wrappingaround junction to a pointattachment junction In that case a “bow up” motion of JC during deformation may disturb or flip the overlaid lipid bilayer and mark stressed erythrocytes for phagocytosisSketches of the erythrocyte membrane skeleton and the wraparound model from large to small scale a The organization of the skeletal network showing JC SC and Sp b A 3D view of a JC with a protofilament associated with three pairs of Sp and 6 SC forming a basic repeating unit27 The angle between JC and the lipid bilayer is not specified c A protofilament having 12 individual G actin and 6 occupied filled binding sites for β Sp d An enlarged section of the protofilament bound with the middle pair Sp3 α3β3 and Sp4 α4β4 Sp3 wraps around a pair of G actin 3a/3b Sp4 wraps around the next pair of G actin 4a/4b in the opposite direction a–d modified from Ref 27 e The detail sketch of α and β spectrin which split and rejoin to wrap the protofilament The tail end begins with the nonhomologous domains of Nβ including ABD CH1 and CH2 and that of Cα including EFs Cys may form disulfide bonds The dimer nucleation site spans α14 and β1821 This sketch appeared in Ref 36 without detailed description The size of linkers of residues Nβ 50 between CH1 and CH2 10 between CH2 and β1 19 between homologous β domains 8 Cα 75 between EF1 and EF2 14 between EFII and α20 8 and between α homologous domains 8 Molecules and domains are not to scale The intertwined feature is not shown for clarity purposesThe αβ Sp is intertwined and arranged in an antiparallel fashion9 In the head region the tetramer is formed with the C terminus of β Sp binding to the N terminus of α Sp which may dissociate into dimers under shear stress In the tail region the N terminus of β Sp is associated with the C terminus of α Sp and binds to the actin protofilament34 On average 6 Sp converge to bind to one actin protofilament in a JC31524 There are 20 homologous domains in α Sp and 16 in β Sp with a 106residue repeat per domain26 The homologous domain pairing near the tail end is responsible for the nucleation for Sp dimerization which is propagated to the head region The initiation of Sp dimerization involves complementary electrostatic interactions between paired triplehelical bundles1 Force extension of α or β Sp by atomic force microscopy revealed a sawtooth pattern that arose from the successive unfolding of the homologous domains each requiring ~30 pN22We previously proposed a wraparound model for the Sp/protofilament junction Fig 1d27 In that model a reinforced protofilament may function as a mechanical axis to anchor three top middle and bottom pairs of Sp Each Sp pair may wraparound the protofilament with a wide dihedral angle ~1662° and a minimal axial distance ~275 nm8 The 3 Sp pairs may spiral down righthanded the protofilament from the pointed end with a dihedral angle of ~554° in between Sp pairs This first 3D model of JC may explain why 6 rather than 12 Sp bind to a protofilament and has been used to simulate the 3D nanomechanics of a single unit33 and multiple units of the membrane skeletal network5 as well as a single unit coupled to the lipid bilayer36 With the support from a number of published biochemical studies we first illustrated how Sp may wraparound the actin protofilament by showing the detailed arrangements of actin domains involved Fig 1d27 followed by detailed arrangements of Sp domains involved Fig 1e36 In this paper we detail the wraparound model in contrast to the pointattachment model generally presented in textbooks or articles6 We first analyze the interaction between intertwined Sp and helical protofilament that begins with one split Sp expands into a pair of Sp and ends with three pairs of Sp We then stepbystep provide an energy and mechanical basis supporting the wraparound model essential for the integrity and elasticity of the erythrocyte membrane skeleton Lastly we show how a wrappingaround junction may be converted to a pointattachment junction leading to the destruction of the network and the death of an erythrocyte This is the novelty of this study advanced from our previous publicationsThe binding affinities were converted into free energy a measure of the reversibility of a reaction ΔG = −RT ln K d where R is the ideal gas constant 83144 J/mol K T is the absolute reference temperature of the system and K d is the binding affinity dissociation constant of the interaction measured in the referenced temperature Thus the binding energy derived from K d can be compared at the same temperature among all interactions involving protofilament α and β Sp and their componentsUsing K B T/p = 8 × 10−21 J/nm and L c = 1442 nm ΔG can be derived The value of K B T/p was derived from the forceextension curve of Sp in the modified WLC model22 where K B is the Boltzmann constant T is the absolute temperature and p is the persistent length of Sp corresponding to 05 nm

Keywords: