[Proteolytic degradation of native hemoglobin and its constituent parts--isolated subunits and globin. I. Kinetic data and the character of the process of the breakdown of native forms].
The rates of proteolytic breakdown for native human hemoglobin (Hb) in CNmet-and oxy-forms, for isolated native alpha- and beta-chains of human Hb with deprotected SH-groups and for apo-Hb--globin at constant temperature 20 degrees as well as for metHb and globin in the temperature range 4-25 degrees were studied. The proteolysis of oxy-forms of proteins was performed in the presence of CN- to prevent the appearance in solution of quickly splitted aqua and hydroxy met-forms. Pepsin (at pH 5.5), trypsin (at pH 7.0 and 8.5) and protease VI (pronase) (at pH 7.0 and 8.5) were used as proteases. The rate of proteolysis was registered simultaneously by proteolysate precipitation in concentrated salt solutions (to determine the content of the native form), by precipitation in aqueous solution of trichloroacetic or perchloric acid and by colouring the terminal NH2-groups by ninhidrin in the total proteolysate. For most cases the data of all the three independent methods fell on a single kinetic curve, each pair protein--protease being represented by their individual curves. Therefore the breakdown of all the protein studied possesses a burst-like ("one-by-one", "all-or-none") character. The protein resistance to the attack by proteolytic enzymes increases in the following order: globin less than oxy-alpha-chain less than metHb less than oxy-beta-chain less than HbO2 congruent to CNmetHb. The use of control repeated proteolysis has made it possible to prove that differences in the rate of proteolytic degradation are not the consequence of spontaneous denaturation of the least unstable forms of proteins in the course of proteolytic reaction but are predetermined by the conformational state of the native macromolecule.
D005914:Globins; D006454:Hemoglobins; D006801:Humans; D007700:Kinetics; D046911:Macromolecular Substances; D010434:Pepsin A / Q000378:metabolism; D011402:Pronase / Q000378:metabolism; D013329:Structure-Activity Relationship; D014357:Trypsin / Q000378:metabolism
D006454:Hemoglobins; D046911:Macromolecular Substances; D005914:Globins; D014357:Trypsin; D010434:Pepsin A; D011402:Pronase
Proteoliticheskaia degradatsiia nativnogo gemoglobina i ego sostavnykh chasteĭ--izolirovannykh sub"edinits i globina. I. Kineticheskie dannye i kharakter protsessa rasshchepleniia nativnykh form.