STRESS RESPONSES - Cyanobacteria - page 655

c) The Clp proteases : A new family of molecular chaperones includes those of caseinolytic proteases
(Clp/Heat shock protein, Hsp100) that consist of constitutive and stress-inducible representatives.
These belong to a broader family of AAA+ proteins (ATPases associated with various cellular
activities) that are known to play different roles such as protein folding, disassembly of protein
complexes and translocation of proteins across membranes. Of the two classes of Clp proteins
known, the fi rst consists of relatively large proteins with two distinct ATP-binding domains (ClpA to
-E and L), while the second has smaller proteins with only one such domain (ClpM, -N, -X, and -Y)
(Thompson and Maurizi, 1994; Kessel et al ., 1995; Schirmer et al ., 1996). The spacer region between
the two ATP-binding domains has been used to classify the Clp genes. ClpA family members
have a spacer of 5 amino acids, ClpC members have 62 to 69 amino acids and ClpB members have
the longest spacer of 123 and 131 amino acids. At least in ClpB, the spacer region of amino acids
comprises 13.5% (Squires and Squires, 1992). ClpA, restricted to gram-negative bacteria like E. coli ,
is the best studied. It is a dimer of 84 kDa subunits and assembles into a hexamer in the presence of
ATP (Maurizi, 1991). It associates with ClpP protein and brings down degradation of the selected
polypeptide. In this process ClpA holds, unfolds and exposes the polypeptide for the proteolytic
activity of ClpP (Hwang et al ., 1987; Katayama-Fujimara et al ., 1987; Thompson and Maurizi, 1994;
Kessel et al ., 1995; Hoskins et al ., 1998; Kim et al ., 1998). Likewise, ClpC and ClpX (but not ClpB)
facilitate the activity of ClpP. ClpA and ClpX exhibit chaperone activity independent of ClpP
analogous to those of DnaK and DnaJ (Wickner et al ., 1994). ClpP proteolytic subunit exhibits low
levels of proteolytic activity but when complexed with ClpA, ClpC or ClpX active holoenzymes are
formed that are able to degrade denatured proteins (Katayama-Fujimara et al ., 1987; Wojtkowlak et al .,
1993; Shanklin et al ., 1995). The proteolytic subunit of ClpP forms a barrel-like structure consisting of
two heptameric rings present opposite to each other. The proteolytc chamber thus formed possesses
the catalytic triad of Ser-His-Asp amino acids. The capping of this chamber on one or both sides
by either ClpA or ClpX hexameric rings takes place (Grimaud et al ., 1998). The interaction of ClpY
with ClpQ to form a proteolytically active holoenzyme has also been reported (Missiakas et al .,
1996). ClpB is another well-characterized member of the Clp/Hsp100 family that is represented
in most eubacteria and eukaryotes. This protein has a long intervening region between the two
ATP-binding domains (Squires and Squires, 1992). Two forms of ClpB proteins are synthesized
in eukaryotes and prokaryotes. In the former, cytosolic-nuclear (100-110 kDa) and mitochondrial
(78 kDa) ClpB proteins are encoded by different genes (Leonhardt et al ., 1993; Sanchez and Lindquist,
1990) but in the latter a single gene encodes a large (94 kDa) and a small (78 kDa) ClpB protein from
a second translational start site (Park et al ., 1993; Eriksson and Clarke, 1996). All ClpB proteins are
induced by high temperatures and most of these are required for thermotolerance. Under normal
conditions ClpB is a non-essential component of cells whose inactivation does not lead to any
phenotypic changes (Sanchez and Lindquist, 1990; Eriksson and Clarke, 1996; Hong and Vierling,
2000). ClpB interacts with large protein aggregates that accumulate under heat stress and the ATP-
induced structural changes enable it to bind to the protein aggregates for performing its function. In
doing so, it cooperates with other Hsps such as DnaK-DnaJ-GrpE to suppress and remove aggregation
of a protein substrate. This has been identifi ed as a highly effi cient multi-chaperone system in E . coli
(Zolkiewski, 1999). The functional collaboration between ClpB and the DnaK system requires the ATP
hydrolysis at the two binding sites of ClpB and these two chaperones act synergistically to remodel
proteins and dissolve aggregates (Doyle et al ., 2007). ClpC is represented in cyanobacteria, plants
and in most gram-positive eubacteria. ClpD is exclusively present in plants. Certain gram-positive
eubacteria possess ClpE and ClpL. ClpB has a protein unfolding activity that is dependent on ATP