Title of article :
Mechanical Unfolding of TNfn3: The Unfolding Pathway of a fnIII Domain Probed by Protein Engineering, AFM and MD Simulation
Author/Authors :
Sean P. Ng، نويسنده , , Ross W.S. Rounsevell، نويسنده , , Annette Steward، نويسنده , , Christian D. Geierhaas، نويسنده , , Philip M. Williams، نويسنده , , Emanuele Paci، نويسنده , , Jane Clarke، نويسنده ,
Issue Information :
روزنامه با شماره پیاپی سال 2005
Pages :
14
From page :
776
To page :
789
Abstract :
Protein engineering Φ-value analysis combined with single molecule atomic force microscopy (AFM) was used to probe the molecular basis for the mechanical stability of TNfn3, the third fibronectin type III domain from human tenascin. This approach has been adopted previously to solve the forced unfolding pathway of a titin immunoglobulin domain, TI I27. TNfn3 and TI I27 are members of different protein superfamilies and have no sequence identity but they have the same β-sandwich structure consisting of two antiparallel β-sheets. TNfn3, however, unfolds at significantly lower forces than TI I27. We compare the response of these proteins to mechanical force. Mutational analysis shows that, as is the case with TI I27, TNfn3 unfolds via a force-stabilised intermediate. The key event in forced unfolding in TI I27 is largely the breaking of hydrogen bonds and hydrophobic interactions between the A′ and G-strands. The mechanical Φ-value analysis and molecular dynamics simulations reported here reveal that significantly more of the TNfn3 molecule contributes to its resistance to force. Both AFM experimental data and molecular dynamics simulations suggest that the rate-limiting step of TNfn3 forced unfolding reflects a transition from the extended early intermediate to an aligned intermediate state. As well as losses of interactions of the A and G-strands and associated loops there are rearrangements throughout the core. As was the case for TI I27, the forced unfolding pathway of TNfn3 is different from that observed in denaturant studies in the absence of force.
Keywords :
AFM , phi-value analysis , tenascin , MD simulation , titin
Journal title :
Journal of Molecular Biology
Serial Year :
2005
Journal title :
Journal of Molecular Biology
Record number :
1245099
Link To Document :
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