A calpain-like proteolytic activity produces the limited cleavage at the N-terminal regulatory domain of rabbit skeletal muscle AMP deaminase: evidence of a protective molecular mechanism.
On storage at 4 degrees C, rabbit skeletal muscle AMP deaminase undergoes limited proteolysis with the conversion of the native 85-kDa enzyme subunit to a 75-kDa core that is resistant to further proteolysis. Further studies have shown that limited proteolysis of AMP deaminase with trypsin, removing the 95-residue N-terminal fragment, converts the native enzyme to a species that exhibits hyperbolic kinetics even at low K+ concentration. The results of this report show that a 21-residue synthetic peptide, when incubated with the purified enzyme, is cleaved with a specificity identical to that reported for ubiquitous calpains. In addition, the cleavage of a specific fluorogenic peptide substrate by rabbit m-calpain is inhibited by a synthetic peptide that corresponds to residues 10-17 of rabbit skeletal muscle AMP deaminase; this peptide contains a sequence (K-E-L-D-D-A) that is present in the fourth subdomain A of rabbit calpastatin, suggesting that the N-terminus of AMP deaminase shares with calpastatin a regulatory sequence that might exert a protective role against the fragmentation-induced activation of AMP deaminase. These observations suggest that a calpain-like proteinase present in muscle removes from AMP deaminase a domain that holds the enzyme in an inactive conformation and which also contains a regulatory region that protects against unregulated proteolysis. We conclude that proteolysis of AMP deaminase is the basis of the large ammonia accumulation that occurs in skeletal muscle subjected to strong tetanic contraction or passing into rigor mortis.
Dipartimento di Scienze dell'Uomo e dell'Ambiente, Chimica e Biochimica Medica, Università di Pisa, Via Roma 55, 56126 Pisa Italy.
D
D
Martini
U
U
Montali
M
M
Ranieri-Raggi
A R M
AR
Sabbatini
S J
SJ
Thorpe
A J G
AJ
Moir
A
A
Raggi
D000659:AMP Deaminase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D002154:Calpain / Q000037:antagonists & inhibitors / Q000378:metabolism*; D004789:Enzyme Activation; D006801:Humans; D018482:Muscle, Skeletal / Q000201:enzymology*; D010455:Peptides / Q000235:genetics / Q000378:metabolism; D011817:Rabbits
D010455:Peptides; D002154:Calpain; D000659:AMP Deaminase