Auto-inhibitory effects of an IQ motif on protein structure and function.
The denuded IQ2 domain, i.e. myosin heavy chain not associated with regulatory light chains, exerts an inhibitory effect on myosin ATPase activity. In this study, we elaborated a structural explanation for this auto-inhibitory effect of IQ2 on myosin function. We employed analytical ultracentrifugation, circular dichroism, and surface plasmon resonance spectroscopy to investigate structural and functional properties of a myosin heavy chain (MYH) head-rod fragment aa664-915. MYH(664-915) was monomeric, adopted a closed shape, and bound essential myosin light chains (HIS-MLC-1) with low affinity to IQ1. Deletion of IQ2, however opened MYH(664-915). Four amino acids present in IQ2 could be identified to be responsible for this auto-inhibitory structural effect: alanine mutagenesis of I814, Q815, R819, and W827 stretched MYH(664-915) and increased 30-fold the binding affinity of HIS-MLC-1 to IQ1. In this study we show, that denuded IQ2 favours a closed conformation of myosin with a low HIS-MLC-1 binding affinity. The collapsed structure of myosin with denuded IQ2 could explain the auto-inhibitory effects of IQ2 on enzymatic activity of myosin.
Petzhold
Daria
D
Max-Delbrück-Center for Molecular Medicine, Department of Molecular Muscle Physiology, Robert-Rössle-Strasse 10, 13125 Berlin, Germany. petzhold@mdc-berlin.de
Lossie
Janine
J
Behlke
Joachim
J
Keller
Sandro
S
Haase
Hannelore
H
Morano
Ingo
I
D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors*; D000818:Animals; D002942:Circular Dichroism; D018995:Myosin Heavy Chains / Q000737:chemistry* / Q000235:genetics; D018994:Myosin Light Chains / Q000737:chemistry; D024462:Myosin Type II / Q000037:antagonists & inhibitors*; D017434:Protein Structure, Tertiary / Q000235:genetics; D051381:Rats; D017384:Sequence Deletion; D020349:Surface Plasmon Resonance; D014461:Ultracentrifugation
D018994:Myosin Light Chains; D000251:Adenosine Triphosphatases; D024462:Myosin Type II; D018995:Myosin Heavy Chains