Mapping residues of SUMO precursors essential in differential maturation by SUMO-specific protease, SENP1.
SUMO (small ubiquitin-related modifier) is a member of the ubiquitin-like protein family that regulates cellular function of a variety of target proteins. SUMO proteins are expressed as their precursor forms. Cleavage of the residues after the 'GG' region of these precursors by SUMO-specific proteases in maturation is a prerequisite for subsequent sumoylation. To understand further this proteolytic processing, we expressed and purified SENP1 (sentrin-specific protease 1), one of the SUMO-specific proteases, using an Escherichia coli expression system. We show that SENP1 is capable of processing all SUMO-1, -2 and -3 in vitro; however, the proteolytic efficiency of SUMO-1 is the highest followed by SUMO-2 and -3. We demonstrate further that the catalytic domain of SENP1 (SENP1C) alone can determine the substrate specificity towards SUMO-1, -2 and -3. Replacement of the C-terminal fragments after the 'GG' region of SUMO-1 and -2 precursors with that of the SUMO-3, indicates that the C-terminal fragment is essential for efficient maturation. In mutagenesis analysis, we further map two residues immediately after the 'GG' region, which determine the differential maturation. Distinct patterns of tissue distribution of SENP1, SUMO-1, -2 and -3 are characterized. Taken together, we suggest that the observed differential maturation process has its physiological significance in the regulation of the sumoylation pathway.
Department of Biochemistry, Faculty of Science, The Chinese University of Hong Kong, Shatin, Hong Kong.
Zheng
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Shannon W N
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D000595:Amino Acid Sequence; D000596:Amino Acids / Q000502:physiology; D020134:Catalytic Domain / Q000502:physiology; D003546:Cysteine Endopeptidases; D010450:Endopeptidases / Q000737:chemistry* / Q000235:genetics; D005786:Gene Expression Regulation / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D009928:Organ Specificity / Q000235:genetics; D010449:Peptide Mapping / Q000379:methods*; D011498:Protein Precursors / Q000737:chemistry; D025842:SUMO-1 Protein; D016415:Sequence Alignment; D025841:Small Ubiquitin-Related Modifier Proteins / Q000737:chemistry* / Q000235:genetics; D013379:Substrate Specificity / Q000502:physiology; D014452:Ubiquitins / Q000737:chemistry / Q000235:genetics
D000596:Amino Acids; D011498:Protein Precursors; D025842:SUMO-1 Protein; C495501:SUMO1 protein, human; C117909:SUMO2 protein, human; C492189:SUMO3 protein, human; D025841:Small Ubiquitin-Related Modifier Proteins; D014452:Ubiquitins; D010450:Endopeptidases; C404512:SENP1 protein, human; D003546:Cysteine Endopeptidases