Glycation of calmodulin: chemistry and structural and functional consequences.
In the presence of Ca2+ and glucose, calmodulin incorporates 2.5 mol of glucose/mol of protein. In the absence of Ca2+, only 1.5 mol of glucose is incorporated per mole of calmodulin. Glycation of calmodulin is associated with variable reductions in its capacity to activate three Ca2+/calmodulin-dependent brain target enzyme systems, including adenylyl cyclase, phosphodiesterase, and protein kinase. In addition, glycated calmodulin exhibits a 54% reduction in its Ca2+ binding capacity. Isolated CNBr cleavage fragments of glycated calmodulin suggest that glycation follows a nonspecific pattern in that each of seven available lysines is susceptible to modification. A limit observed on the extent of glycation appears related to the accompanying increase in negative charge on the protein. Glycation results in minimal structural rearrangements in calmodulin, and the Ca2+-induced increase in alpha-helix content and radius of gyration is the same for glycated and unmodified calmodulin. Since glycated calmodulin's Ca2+ binding capacity is reduced, this implies that the Ca2+-induced conformational changes in calmodulin do not require all four Ca2+ binding sites to be occupied. Examination of the lysine positions in calmodulin suggests that Ca2+ binding to domains II and IV is sufficient to induce these changes. The functional consequences of calmodulin glycation therefore cannot be attributed to inhibition of these conformational changes. An alternative explanation is that the inhibition arises from interference at the target enzyme binding site by bound glucose. While glycation shows minimal structural effects, a large pH dependence is observed for the alpha-helix content of unmodified calmodulin.(ABSTRACT TRUNCATED AT 250 WORDS)
Kowluru
R A
RA
Life Sciences and Physics Divisions, Los Alamos National Laboratory, New Mexico 87545.
Heidorn
D B
DB
Edmondson
S P
SP
Bitensky
M W
MW
Kowluru
A
A
Downer
N W
NW
Whaley
T W
TW
Trewhella
J
J
D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D001923:Brain Chemistry; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism*; D002417:Cattle; D002942:Circular Dichroism; D003488:Cyanogen Bromide; D004789:Enzyme Activation; D006031:Glycosylation; D066298:In Vitro Techniques; D008239:Lysine / Q000378:metabolism; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D011494:Protein Kinases / Q000378:metabolism; D012542:Scattering, Radiation; D014965:X-Rays
D002147:Calmodulin; D011494:Protein Kinases; D010727:Phosphoric Diester Hydrolases; D000262:Adenylyl Cyclases; D008239:Lysine; D003488:Cyanogen Bromide; D002118:Calcium