The cytotoxin alpha-sarcin behaves as a cyclizing ribonuclease.
The hydrolysis of adenylyl(3'-->5')adenosine (ApA) and guanylyl(3'--> 5')adenosine (GpA) dinucleotides by the cytotoxic protein alpha-sarcin has been studied. Quantitative analysis of the reaction has been performed through reverse-phase chromatographic (HPLC) separation of the resulting products. The hydrolysis of the 3'-5' phosphodiester bond of these substrates yields the 2'-3' cyclic mononucleotide; this intermediate is converted into the corresponding 3'-monophosphate derivative as the final product of the reaction. The values of the apparent Michaelis constant (KM), kcat and kcat/KM have also been calculated. The obtained results fit into a two-step mechanism for the enzymatic activity of alpha-sarcin and allow to consider this protein as a cyclizing RNase.
Lacadena
J
J
Departamento de Bioquímica y Biología Molecular I, Facultad de Química, Universidad Complutense de Madrid, Spain.
Martínez del Pozo
A
A
Lacadena
V
V
Martínez-Ruiz
A
A
Mancheño
J M
JM
Oñaderra
M
M
Gavilanes
J G
JG
D001230:Aspergillus / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D004722:Endoribonucleases / Q000302:isolation & purification / Q000378:metabolism*; D005656:Fungal Proteins; D006868:Hydrolysis; D009843:Oligoribonucleotides / Q000378:metabolism*; D012260:Ribonucleases / Q000378:metabolism*
D005656:Fungal Proteins; D009843:Oligoribonucleotides; C014801:alpha-sarcin; D004722:Endoribonucleases; D012260:Ribonucleases