Maleimidobenzoyl-G-actin: structural properties and interaction with skeletal myosin subfragment-1.
We have investigated various structural and interaction properties of maleimidobenzoyl-G-actin (MBS-actin), a new, internally cross-linked G-actin derivative that does not exhibit, at moderate protein concentration, the salt--and myosin subfragment 1 (S-1)-induced polymerizations of G-actin and reacts reversibly and covalently in solution with S-1 at or near the F-actin binding region of the heavy chain (Bettache, N., Bertrand, R., & Kassab, R. (1989) Proc. Natl. Acad. Sci. U.S.A. 86, 6028-6032). The far-ultraviolet CD spectrum and alpha-helix content of the MBS-actin were identical with those displayed by native G-actin. 45Ca2+ measurements showed the same content of tightly bound Ca2+ in MBS-actin as in G-actin and the EDTA treatment of the modified protein promoted the same red shift of the intrinsic fluorescence spectrum as observed with native G-actin. Incubation of concentrated MBS-actin solutions with 100 mM KCl + 5 mM MgCl2 led to the polymerization of the actin derivative when the critical monomer concentration reached 1.6 mg/mL, at 25 degrees C, pH 8.0. The MBS-F-actin formed activated the Mg2(+)-ATPase of S-1 to the same extent as native F-actin. The MBS-G-actin exhibited a DNase I inhibitor activity very close to that found with native G-actin and was not to be at all affected by its specific covalent conjugation to S-1.(ABSTRACT TRUNCATED AT 250 WORDS)
Bettache
N
N
Centre de Recherches de Biochimie Macromoléculaire du CNRS, INSERM U249, Université de Montpellier I, France.
Bertrand
R
R
Kassab
R
R
D000199:Actins / Q000737:chemistry* / Q000378:metabolism; D000818:Animals; D002942:Circular Dichroism; D003432:Cross-Linking Reagents / Q000737:chemistry*; D003850:Deoxyribonuclease I / Q000378:metabolism*; D007700:Kinetics; D015879:Myosin Subfragments / Q000737:chemistry* / Q000378:metabolism; D011487:Protein Conformation; D011817:Rabbits; D013050:Spectrometry, Fluorescence; D013329:Structure-Activity Relationship; D014357:Trypsin
D000199:Actins; D003432:Cross-Linking Reagents; D015879:Myosin Subfragments; C066969:maleimidobenzoyl-G-actin; D003850:Deoxyribonuclease I; D014357:Trypsin