Description:
<jats:p>The composition of the chloroplast‐localized protease complex, ClpP, from the green alga <jats:italic>Chlamydomonas reinhardtii</jats:italic> was characterized by nondenaturing electrophoresis, immunoblotting and MS. The detected ClpP complex has a native mass of ≈ 540 kDa, which is ≈ 200 kDa higher than ClpP complexes in higher plant chloroplasts, mitochondria or bacteria. The 540‐kDa ClpP complex contains two nuclear‐encoded ClpP proteins (ClpP3 and P5) and five ClpR (R1, R2, R3, R4 and R6) proteins, as well two proteins, ClpP1<jats:sub>L</jats:sub> and ClpP1<jats:sub>H</jats:sub>, both probably derived from the plastid <jats:italic>clpP1</jats:italic> gene. ClpP1<jats:sub>H</jats:sub> is 59 kDa and contains a ≈ 30‐kDa insertion sequence (IS1) not found in other ClpP proteins, responsible for the high MW of the complex. Based on comparison with other sequences, IS1 protrudes as an additional domain on the apical surface of the ClpP/R complex, probably preventing interaction with the HSP100 chaperone. ClpP1<jats:sub>L</jats:sub> is a 25‐kDa protein similar in size to other ClpP proteins and could arise by post‐translational processing of ClpP1<jats:sub>H</jats:sub>. Chloramphenicol‐chase experiments show that ClpP1<jats:sub>L</jats:sub> and ClpP1<jats:sub>H</jats:sub> have a similar half‐life, indicating that both are stable components of the complex. The structure of the ClpP complex is further discussed in conjunction with a phylogenetic analysis of the ClpP/R genes. A model is proposed for the evolution of the algal and plant complex from its cyanobacterial ancestor.</jats:p>