The calcium-induced switch in the troponin complex probed by fluorescent mutants of troponin I.
The Ca2+-induced transition in the troponin complex (Tn) regulates vertebrate striated muscle contraction. Tn was reconstituted with recombinant forms of troponin I (TnI) containing a single intrinsic 5-hydroxytryptophan (5HW). Fluorescence analysis of these mutants of TnI demonstrate that the regions in TnI that respond to Ca2+ binding to the regulatory N-domain of TnC are the inhibitory region (residues 96-116) and a neighboring region that includes position 121. Our data confirms the role of TnI as a modulator of the Ca2+ affinity of TnC; we show that point mutations and incorporation of 5HW in TnI can affect both the affinity and the cooperativity of Ca2+ binding to TnC. We also discuss the possibility that the regulatory sites in the N-terminal domain of TnC might be the high affinity Ca2+-binding sites in the troponin complex.
Oliveira
Deodoro C S G
DC
Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, Brazil.
Reinach
Fernando C
FC
D006916:5-Hydroxytryptophan / Q000737:chemistry / Q000378:metabolism; D000494:Allosteric Regulation; D001665:Binding Sites; D002118:Calcium / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D017354:Point Mutation / Q000502:physiology; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D014335:Tropomyosin / Q000378:metabolism / Q000502:physiology*; D014336:Troponin / Q000378:metabolism / Q000502:physiology*; D019210:Troponin I / Q000235:genetics / Q000378:metabolism / Q000502:physiology*
D011994:Recombinant Proteins; D014335:Tropomyosin; D014336:Troponin; D019210:Troponin I; C042260:troponin-tropomyosin complex; D006916:5-Hydroxytryptophan; D002118:Calcium