title: Biochemical characterization of the small ubiquitin-like modifiers of Chlamydomonas reinhardtii.
abstract: Dynamic modification of target proteins by small ubiquitin-like modifier (SUMO) is known to modulate many important cellular processes and is required for cell viability and development in all eukaryotes. However, understanding of SUMO systems in plants, especially in unicellular green algae, remains elusive. In this study, Chlamydomonas reinhardtii CrSUMO96, CrSUMO97 and CrSUMO148 were characterized. We show that the formation of polymeric CrSUMO96 and CrSUMO97 chains can be catalyzed either by the human SAE1/SAE2 and Ubc9 SUMOylation system in vitro or by an Escherichia coli chimeric SUMOylation system in vivo. An exposed C-terminal di-glycine motif of CrSUMO96 or CrSUMO97 is essential for functional SUMOylation. The human SUMO-specific protease, SENP1, demonstrates more processing activity for CrSUMO97 than for CrSUMO96. The CrSUMO148 precursor notably has four repeated di-glycine motifs at the C-terminus. This unique feature is not found in other known SUMO proteins. Interestingly, only 83-residual CrSUMO148(1-83) with the first di-glycine motif can form SAE1/SAE2-SUMO complex and further form polymeric chains with the help of Ubc9. More surprisingly, CrSUMO148 precursor is digested by SENP1, solely at the peptide bond after the first di-glycine motif although there are four theoretically identical processing sites in the primary sequence. This process directly generates 83-residual CrSUMO148(1-83) mature protein, which is exactly the form suitable for activation and conjugation. We also show that SENP1 displays similar isopeptidase activity in the deconjugation of polymeric CrSUMO96, CrSUMO97 or CrSUMO148 chains, revealing that the catalytic mechanisms of processing and deconjugation of CrSUMOs by SENP1 may differ.
authors.affiliation: Department of Biochemical Science and Technology, Institute of Microbiology and Biochemistry, College of Life Science, National Taiwan University, No 1, Sec 4, Roosevelt Road, Taipei 10617, Taiwan.
authors.forename: Yung-Cheng
authors.initials: YC
authors.lastname: Shin
authors.forename: Bang-Yu
authors.initials: BY
authors.lastname: Liu
authors.forename: Jia-Yun
authors.initials: JY
authors.lastname: Tsai
authors.forename: Jiunn-Tzong
authors.initials: JT
authors.lastname: Wu
authors.forename: Li-Kwan
authors.initials: LK
authors.lastname: Chang
authors.forename: Shih-Chung
authors.initials: SC
authors.lastname: Chang
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D016825:Chlamydomonas reinhardtii / Q000235:genetics / Q000378:metabolism*; D017931:DNA Primers; D008958:Models, Molecular; D008969:Molecular Sequence Data; D018749:RNA, Plant / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D025841:Small Ubiquitin-Related Modifier Proteins / Q000737:chemistry / Q000378:metabolism; D025801:Ubiquitin / Q000378:metabolism*
chemical_list: D017931:DNA Primers; D018749:RNA, Plant; D025841:Small Ubiquitin-Related Modifier Proteins; D025801:Ubiquitin