Stopped-flow kinetic studies of Ca(II) and Mg(II) dissociation in cod parvalbumin and bovine alpha-lactalbumin.
The dissociation kinetics of complexes of bovine alpha-lactalbumin and cod parvalbumin with Ca(II) and Mg(II) ions induced by mixing of a Ca(II)- or MG(II)-loaded protein with a chelator of divalent cations (EDTA or EGTA) have been studied by means of the stopped-flow method with intrinsic protein fluorescence registration. Within the temperature interval from 10 to approx. 37 degrees C kinetic curves for Ca(II) removal from alpha-lactalbumin are monoexponential with a rate constant ranging from 0.006 to 1 s. Taking into account the rather low rate of fluorescence changes, one can assume that the limiting stage in this case is the dissociation of the single bound Ca(II) ion from the protein and not a conformational transition which occurs after Ca(II) dissociation. At temperatures above 37 degrees C the kinetic curves require at least two exponential terms for a satisfactory fit. The second exponential seems to be due to denaturation of the apo form of alpha-lactalbumin which takes place at these temperatures. The values of the dissociation rate constants for Mg(II) bound to alpha-lactalbumin practically coincide with those for Ca(II). Within the temperature interval 10-30 degrees C the kinetic curves for Ca(II) and Mg(II) removal from parvalbumin are best fitted by a sum of two exponential terms identified as arising from the dissociation of cations from the two binding sites.(ABSTRACT TRUNCATED AT 250 WORDS)
Institute of Biological Physics of the U.S.S.R. Academy of Sciences, Pushchino, Moscow Region.
E A
EA
Permyakov
A V
AV
Ostrovsky
L P
LP
Kalinichenko
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002417:Cattle; D005399:Fishes; D007700:Kinetics; D007768:Lactalbumin / Q000378:metabolism*; D008274:Magnesium / Q000378:metabolism*; D009124:Muscle Proteins / Q000378:metabolism*; D010320:Parvalbumins / Q000378:metabolism*; D011485:Protein Binding; D013816:Thermodynamics
D009124:Muscle Proteins; D010320:Parvalbumins; D007768:Lactalbumin; D008274:Magnesium; D002118:Calcium