Global shapes of F-actin depolymerization-competent minimal gelsolins: insight into the role of g2-g3 linker in pH/Ca2+ insensitivity of the first half.
Because of its ability to rapidly depolymerize F-actin, plasma gelsolin has emerged as a therapeutic molecule in different disease conditions. High amounts of exogenous gelsolin are, however, required to treat animal models of different diseases. Knowing that the F-actin depolymerizing property of gelsolin resides in its N terminus, we made several truncated versions of plasma gelsolin. The smaller versions, particularly the one composed of the first 28-161 residues, depolymerized the F-actin much faster than the native gelsolin and other truncates at the same molar ratios. Although G1-G3 loses its dependence on Ca(2+) or low pH for the actin depolymerization function, interestingly, G1-G2 and its smaller versions were found to regain this requirement. Small angle x-ray scattering-based shape reconstructions revealed that G1-G3 adopts an open shape in both the presence and the absence of Ca(2+) as well as low pH, whereas G1-G2 and residues 28-161 prefer collapsed states in Ca(2+)-free conditions at pH 8. The mutations in the g2-g3 linker resulted in the calcium sensitivity of the mutant G1-G3 for F-actin depolymerization activity, although the F-actin-binding sites remained exposed in the mutant G1-G3 as well as in the smaller truncates even in the Ca(2+)-free conditions at pH 8. Furthermore, unlike wild type G1-G3, calcium-sensitive mutants of G1-G3 acquired closed shapes in the absence of free calcium, implying a role of g2-g3 linker in determining the open F-actin depolymerizing-competent shape of G1-G3 in this condition. We demonstrate that the mobility of the G1 domain, essential for F-actin depolymerization, is indirectly regulated by the gelsolin-like sequence of g2-g3 linker.
From the Council of Scientific and Industrial Research-Institute of Microbial Technology, Chandigarh 160036, India.
Nagesh
N
Peddada
Amin
A
Sagar
Yogendra S
YS
Rathore
Vikas
V
Choudhary
U Bharat K
UB
Pattnaik
Neeraj
N
Khatri
Renu
R
Garg
Ashish
D000199:Actins / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002645:Chickens; D003001:Cloning, Molecular; D018260:Gelsolin / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018613:Microscopy, Confocal; D008958:Models, Molecular; D009132:Muscles / Q000378:metabolism; D009154:Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012542:Scattering, Radiation; D018805:Sepsis / Q000378:metabolism; D014965:X-Rays
D000199:Actins; D018260:Gelsolin; D002118:Calcium
Actin; Minimal Gelsolin; Molecular Modeling; Protein Engineering; Sepsis; Small Angle X-ray Scattering; X-ray Scattering; ab Initio Modeling