2'-Phosphoadenylylation of eukaryotic proteins: a type of covalent modification.
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2'-Phosphoadenylylation of eukaryotic proteins: a type of covalent modification. / Hilz, H; Fanick, Werner; Klapproth, K.
In: P NATL ACAD SCI USA, Vol. 83, No. 17, 17, 1986, p. 6267-6271.Research output: SCORING: Contribution to journal › SCORING: Journal article › Research › peer-review
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TY - JOUR
T1 - 2'-Phosphoadenylylation of eukaryotic proteins: a type of covalent modification.
AU - Hilz, H
AU - Fanick, Werner
AU - Klapproth, K
PY - 1986
Y1 - 1986
N2 - An enzymatic system in rat liver microsomal preparations has been detected that catalyzes the transfer of the 2'-phospho-AMP moiety from NADP to endogenous polypeptides; the major acceptor is a polypeptide of about 40 kDa (p40). Modification of the acceptor by 2'-phospho-AMP residues was deduced from the simultaneous transfer of 2'-[33P]phosphate and [3H]adenine residues from double-labeled NADP, while no incorporation of radioactivity into p40 was seen with NADP species labeled in the NMN moiety. The true substrate of this phosphoadenylylation reaction was 2'-phospho-ADP-ribose rather than NADP, because labeled phospho-ADP-ribose was as efficient as or more efficient than NADP in forming modified p40. Also, NADP was rapidly converted to phospho-ADP-ribose during incubation with microsomes. Furthermore, isonicotinic acid hydrazide, an inhibitor of NADP glycohydrolase, prevented phosphoadenylylation from NADP, but not from phospho-ADP-ribose, and glycohydrolase-resistant NADPH could not substitute for NADP. Transferase activity was found in liver and brain microsomes and, to a smaller extent, in the cytosol fractions. In Ehrlich ascites tumor cells, most of the activity resided in the cytosol, from which it could be partially purified. The apparent Km for phospho-ADP-ribose was about 2 X 10(-4) M, and the pH optimum was around 7. Divalent cations like Mg2+ and Mn2+ inhibited the reaction. In all compartmental preparations, activity was eliminated by heating or short treatment with alkali or acid. In submitochondrial particles from rat liver, a system with different characteristics led to the phosphoadenylylation of several endogenous polypeptides.
AB - An enzymatic system in rat liver microsomal preparations has been detected that catalyzes the transfer of the 2'-phospho-AMP moiety from NADP to endogenous polypeptides; the major acceptor is a polypeptide of about 40 kDa (p40). Modification of the acceptor by 2'-phospho-AMP residues was deduced from the simultaneous transfer of 2'-[33P]phosphate and [3H]adenine residues from double-labeled NADP, while no incorporation of radioactivity into p40 was seen with NADP species labeled in the NMN moiety. The true substrate of this phosphoadenylylation reaction was 2'-phospho-ADP-ribose rather than NADP, because labeled phospho-ADP-ribose was as efficient as or more efficient than NADP in forming modified p40. Also, NADP was rapidly converted to phospho-ADP-ribose during incubation with microsomes. Furthermore, isonicotinic acid hydrazide, an inhibitor of NADP glycohydrolase, prevented phosphoadenylylation from NADP, but not from phospho-ADP-ribose, and glycohydrolase-resistant NADPH could not substitute for NADP. Transferase activity was found in liver and brain microsomes and, to a smaller extent, in the cytosol fractions. In Ehrlich ascites tumor cells, most of the activity resided in the cytosol, from which it could be partially purified. The apparent Km for phospho-ADP-ribose was about 2 X 10(-4) M, and the pH optimum was around 7. Divalent cations like Mg2+ and Mn2+ inhibited the reaction. In all compartmental preparations, activity was eliminated by heating or short treatment with alkali or acid. In submitochondrial particles from rat liver, a system with different characteristics led to the phosphoadenylylation of several endogenous polypeptides.
KW - Animals
KW - Rats
KW - Molecular Weight
KW - Protein Processing, Post-Translational
KW - Phosphoproteins/metabolism
KW - Adenosine Diphosphate Ribose/analogs & derivatives/metabolism
KW - Adenosine Monophosphate/analogs & derivatives/metabolism
KW - Cell Compartmentation
KW - Chemical Phenomena
KW - Chemistry
KW - Microsomes, Liver/metabolism
KW - NADP/metabolism
KW - Nucleoside Diphosphate Sugars
KW - Animals
KW - Rats
KW - Molecular Weight
KW - Protein Processing, Post-Translational
KW - Phosphoproteins/metabolism
KW - Adenosine Diphosphate Ribose/analogs & derivatives/metabolism
KW - Adenosine Monophosphate/analogs & derivatives/metabolism
KW - Cell Compartmentation
KW - Chemical Phenomena
KW - Chemistry
KW - Microsomes, Liver/metabolism
KW - NADP/metabolism
KW - Nucleoside Diphosphate Sugars
M3 - SCORING: Journal article
VL - 83
SP - 6267
EP - 6271
JO - P NATL ACAD SCI USA
JF - P NATL ACAD SCI USA
SN - 0027-8424
IS - 17
M1 - 17
ER -
