α‑Helical Domains Affecting the Oligomerization of Vipp1 and Its Interaction with Hsp70/DnaK in Chlamydomonas

• Published in: Biochemistry (Easton) 2015-08, Vol.54 (31), p.4877-4889
• Main Authors: Gao, Fei, Wang, Wenyan, Zhang, Wenjuan, Liu, Cuimin
• Format: Article
• Language: English
• Subjects: Chlamydomonas; Chlamydomonas reinhardtii - chemistry; Chlamydomonas reinhardtii - genetics; Chlamydomonas reinhardtii - metabolism; Chloroplast Proteins - chemistry; Chloroplast Proteins - genetics; Chloroplast Proteins - metabolism; HSP70 Heat-Shock Proteins - chemistry; HSP70 Heat-Shock Proteins - genetics; HSP70 Heat-Shock Proteins - metabolism; Membrane Proteins - chemistry; Membrane Proteins - genetics; Membrane Proteins - metabolism; Protein Multimerization - physiology; Protein Structure, Quaternary; Protein Structure, Secondary; Protein Structure, Tertiary
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/acs.biochem.5b00050

Vesicle-inducing protein in plastids 1 (Vipp1) forms >1 MDa ordered homo-oligomeric complexes in chloroplasts and is involved in the biogenesis of photosynthetic machinery. The Hsp70 chaperone system has been shown to interact with Vipp1, influencing its higher-order structure. In this study, a series of deletion mutants in Chlamydomonas reinhardii Vipp1 (CrVipp1) is used to investigate the role of the α-helical domains (H1–H7) in mediating its structure and interaction with DnaK, an Hsp70 orthologue. Results from these analyses demonstrate that α-helical domains H1–H6 of CrVipp1 are required for its efficient accumulation in protease-resistant large complexes, termed superoligomers. Deletions of these α-helical domains, either individually or in combination, cause CrVipp1 to assemble into a heterogeneous mixture of smaller, protease-sensitive oligomers. Furthermore, domains H2 and H3 are required to form a stable structural core in mutant oligomers, whereas domains H1 and H4–H6 likely function downstream in assembly of the superoligomer. DnaK binds only weakly to any form of CrVipp1 that efficiently assembles into superoligomers. In contrast, the interaction with DnaK is much more robust with certain misfolded CrVipp1 oligomers in a process mediated by their H4 and H7 domains. DnaK also interacts with full-length CrVipp1 at an early stage of CrVipp1 biosynthesis, perhaps during initial steps in the oligomerization pathway. Taken together, these data suggest that not only α-helical domains but also the oligomeric states of CrVipp1 influence its interaction with DnaK. It is therefore plausible that the Hsp70/DnaK system may be involved in the assembly of Vipp1 superoligomers.