Asymmetric functional interaction between chaperonin and its plastidic cofactors

Abstract

The specific cochaperonin, chloroplast chaperonin (Cpn)20, consisting of two tandem GroES-like domains, is present abundantly in plant and algal chloroplasts, in addition to Cpn10, which is similar in size to GroES. How Cpn20 oligomers, containing six or eight 10-kDa domains, cooperate with the heptameric ring of chaperonin at the same time as encountering symmetry mismatch is unclear. In the present study, we characterized the functional cooperation of cochaperonins, including two plastidic Cpn20 homo-oligomers from Arabidopsis (AtCpn20) and Chlamydomonas (CrCPN20), and one algal CrCPNs hetero-oligomer, consisting of three cochaperonins, CrCPN11, CrCPN20 and CrCPN23, with two chaperonins, Escherichia coli GroEL and Chlamydomonas CrCPN60. AtCpn20 and CrCPNs were functional for assisting both chaperonins in folding model substrates ribulose bisphosphate carboxylase oxygenase from Rhodospirillum rubrum (RrRubisco) in vitro and complementing GroES function in E. coli. CrCPN20 cooperated only with CrCPN60 (and not GroEL) to refold RrRubisco in vitro and showed differential complementation with the two chaperonins in E. coli. Cochaperonin concatamers, consisting of six to eight covalently linked 10-kDa domains, were functionally similar to their respective native forms. Our results indicate that symmetrical match between chaperonin and cochaperonin is not an absolute requisite for functional cooperation.

Hetero‐oligomeric CPN60 resembles highly symmetric group‐I chaperonin structure revealed by Cryo‐EM

“…Rotational correlation analysis of CPN11/20/23 revealed high similarity among the GroES‐like domains of the different co‐chaperonin subunits (Figure a,b). Hence, the CPN11/20/23 complex matches well with the heptameric cis ‐ring of CPN60α/β1/β2, therefore solving the symmetry problem by co‐chaperonin subunit hetero‐oligomerization (Weiss et al ., ; Tsai et al ., ; Vitlin Gruber et al ., ; Guo et al ., ).…”

“…The Rubisco carboxylase activity assay was performed as described previously (Guo et al ., ). Samples containing refolded Rubisco were mixed with 15 m m NaHCO 3 , 0.1 μCi per μl NaH 14 CO 3 , 20 m m MgCl 2 and 0.2 m m DTT, and then incubated for 5 min.…”

“…Furthermore, domain swapping between CPN60a and CPN60b demonstrated that the CPN60a apical domain could not functionally cooperate with co-chaperonin GroES, but recognized its cognate substrate, the Rubisco large subunit from Chlamydomonas, more efficiently than the CPN60b apical domain, and vice versa, suggesting functional differentiation (Zhang et al, 2016b). Regarding chloroplast co-chaperonins, it is not clear how the six-or eightfold symmetry realized in functional Cpn20 homo-oligomers matches with the heptameric chaperonin cylinder, although a recent biochemical study has claimed that a perfect match with the chaperonin was no absolute prerequisite for a functional interaction (Baneyx et al, 1995;Koumoto et al, 1999;Guo et al, 2015). Several in vitro studies have suggested that, similar to Cpn60s, the co-chaperonins might also form Cpn20-Cpn10 hetero-oligomers in vivo (Tsai et al, 2012;Vitlin Gruber et al, 2014).…”

Hetero-oligomeric CPN60 resembles highly symmetric group I chaperonin structure revealed by Cryo-EM

“…Furthermore, domain swappings between CPN60α and CPN60β demonstrated that the CPN60α apical domain could not functionally cooperate with co-chaperonin GroES, but recognized its cognate substrate, the Rubisco large subunit from Chlamydomonas, more efficiently than the CPN60β apical domain and vice versa, suggesting one way of functional differentiation (Zhang et al, 2016b). Regarding chloroplast co-chaperonins it is not clear how the six/eight-fold symmetry realized in functional Cpn20 homo-oligomers matches with the heptameric chaperonin cylinder, although a recent biochemical study has claimed that a prefect match with the chaperonin was no absolute prerequisite for a functional interaction (Baneyx et al, 1995;Guo et al, 2015;Koumoto et al, 1999). Several in vitro studies have suggested that, similar to Cpn60s, the co-chaperonins might also form Cpn20-Cpn10 hetero-oligomers in vivo (Tsai et al, 2012;Vitlin Gruber et al, 2014).…”

Non‐housekeeping, non‐essential GroEL (chaperonin) has acquired novel structure and function beneficial under stress in cyanobacteria

“…In C. reinhardtii , there are also three co‐Cpn genes, with one gene encoding Cpn10 (Cpn11) and the other two encoding Cpn20 and Cpn23 with tandem Cpn10 sequences (Tsai et al ). Cpn10 and Cpn20/Cpn23 from A. thaliana and C. reinhardtii can assemble into functional homo‐oligomeric and/or hetero‐oligomeric complexes in vitro and/or in vivo (in an E. coli cell) (Tsai et al , Gruber et al , Guo et al ). This heterogeneity may confer chloroplast Cpn60s and their co‐Cpns great diversity in their structures and functions which may be required specifically for chloroplast development and function.…”