Chemical modification of glycerinated stalks shows tyrosine residues essential for spasmoneme contraction of Vorticella sp.
Chemical modification of glycerinated stalks of Vorticella with TNM is used to investigate the role of tyrosine residues in the Ca(2+)-induced contraction of the spasmoneme. Tetranitromethane (TNM) is often employed as a specific reagent for the nitration of tyrosine residues in a protein at neutral and slightly alkaline pHs although TNM can also oxidize cysteine residues in the acidic and neutral pH range. Prior incubation with Ca(2+) of stalks to be treated with TNM can protect the spasmoneme from irreversible denaturation. On the other hand, TNM treatment in the absence of free Ca(2+) causes an irreversible denaturation of the spasmoneme. It was revealed by us that an isolated Ca(2+)-binding protein called spasmin could not bind with Ca(2+) after TNM treatment, even if the treatment was performed in the presence of Ca(2+). In an additional experiment, we confirmed that the chemical modification of cysteine residues in the spasmoneme with N-7-dimethyl-amino-4methyl- coumarinyl- maleimide (DACM) has no effect on the contractibility. These results suggest that tyrosine residues in spasmin are essential for spasmoneme contraction and are protected from TNM in the presence of Ca(2+) when spasmin binds with its receptor protein in the spasmoneme.
Fang
Jie
J
Department of Bioscience and Biomedical Engineering, Graduate School of Science and Engineering, Waseda University, Tokyo, Japan.
Zhang
Bei
B
Chen
Ning
N
Asai
Hiroshi
H
D000825:Animal Structures / Q000187:drug effects* / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002681:China; D003285:Contractile Proteins / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D008301:Maleimides / Q000378:metabolism / Q000494:pharmacology; D016802:Oligohymenophorea / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects; D011489:Protein Denaturation / Q000187:drug effects; D015800:Protozoan Proteins / Q000378:metabolism*; D013774:Tetranitromethane / Q000378:metabolism / Q000494:pharmacology*; D014443:Tyrosine / Q000378:metabolism
D003285:Contractile Proteins; D008301:Maleimides; D015800:Protozoan Proteins; C026555:spasmin; D014443:Tyrosine; C019347:N-(7-dimethylamino-4-methylcoumarinyl)maleimide; D013774:Tetranitromethane; D002118:Calcium