FTSH (HFLB) IS AN ATP-DEPENDENT PROTEASE SELECTIVELY ACTING ON SECY AND SOME OTHER MEMBRANE-PROTEINS

The FtsH protein is a membrane-bound ATPase of Escherichia coil that w as proposed to be involved in membrane protein assembly as well as deg radation of some unstable proteins, SecY, a subunit of protein translo case, is FtsH dependently degraded in vivo when it fails to associate with its partner (the SecE protein), We constructed a series of mutant s in which mutations were introduced into conserved residues in the tw o ATP binding consensus sequences or the zinc binding sequence of FtsH , We purified wild-type and mutant FtsH proteins by making use of a po lyhistidine tag attached to their carboxyl termini, Complementation an alysis and ATPase activity assays in vitro indicated that, of the two sets of ATP binding sequence motifs, the one located C-terminally (Al) is essential for ATPase activity and in two functioning of FtsH, Wild -type FtsH protein degraded purified SecY in an ATP hydrolysis-depende nt manner in vitro. Mutant proteins without ATPase activity were inact ive in proteolysis. A zinc binding motif mutant showed a decreased pro teolytic activity. SecY and FtsH were cross linkable with each other i n the membrane, provided that FtsH had an ATPase-inactivating mutation , These results demonstrate that FtsH binds to and degrades SecY, its Al motif and the zinc binding motif being important for the proteolyti c activity. FtsH-dependent proteolysis was also demonstrated for SecY in crude membrane extracts, whereas a majority of other membrane prote ins were not degraded, indicating that FtsH has high selectivity in pr otein degradation.