Yoshinori Akiyama 교수

Escherichia coli YaeL (EcfE) is a homolog of human site-2 protease (S2P), a membrane-bound zinc metalloprotease involved in regulated intramembrane proteolysis. We have shown previously that YaeL, having essential metalloprotease active site motifs in the cytoplasmic domain, is indispensable for viability. Here, we obtained rpoE, encoding an extracytoplasmic stress response sigma factor (sigma(E)), as a multicopy suppressor against the yaeL disruption. Whereas sigma(E) is thought to be activated

The FtsH protein is a membrane-bound ATPase of Escherichia coli that was proposed to be involved in membrane protein assembly as well as degradation of some unstable proteins. SecY, a subunit of protein translocase, is FtsH dependently degraded in vivo when it fails to associate with its partner (the SecE protein). We constructed a series of mutants in which mutations were introduced into conserved residues in the two ATP binding consensus sequences or the zinc binding sequence of FtsH. We purif

Escherichia coli FtsH is a membrane-bound ATPase with a proteolytic activity against the SecY subunit of protein translocase. We now report that subunit a of the membrane-embedded Fo part of H+-ATPase is another substrate of FtsH. Pulse-chase experiments showed that subunit a is unstable when it alone (without Fo subunits b and c) was oversynthesized and that it is stabilized in the ftsH mutants. Selective and ATP-dependent degradation of subunit a by purified FtsH protein was demonstrated in vi

It was shown previously that the Escherichia coli gene ppfA (dsbA) encodes a periplasmic protein, and its inactivation leads to a deficiency in disulfide bond formation of envelope proteins (Kamitani, S., Akiyama, Y., and Ito, K. (1992) EMBO J. 11, 57-62; Bardwell, J. C. A., McGovern, K., and Beckwith, J. (1991) Cell 67, 581-589). The DsbA/PpfA protein was overproduced, purified, and examined for its activities in vitro. Its abundance in a wild-type cell was estimated to be about 850 molecules w

Taken together, these results suggest that the Cpx "extracytoplasmic" stress response system controls the quality of the plasma membrane, even on its cytoplasmic side.

Escherichia coli HtpX is a putative membrane-bound zinc metalloprotease that has been suggested to participate in the proteolytic quality control of membrane proteins in conjunction with FtsH, a membrane-bound and ATP-dependent protease. Here, we biochemically characterized HtpX and confirmed its proteolytic activities against membrane and soluble proteins. HtpX underwent self-degradation upon cell disruption or membrane solubilization. Consequently, we purified HtpX under denaturing conditions

The FtsH (HflB) protein of Escherichia coli is integrated into the membrane with two N-terminally located transmembrane segments, while its large cytoplasmic domain is homologous to the AAA family of ATPases. The previous studies on dominant negative ftsH mutants raised a possibility that FtsH functions in multimeric states. We found that FtsH was eluted at fractions corresponding to a larger molecular weight than expected from monomeric structure in size-exclusion chromatography. Moreover, trea

FtsH is an Escherichia coli protein with its amino-terminal region anchored to the cytoplasmic membrane and with its cytoplasmic domain significantly homologous to the members of an ATPase family found in eukaryotic cells. We previously showed that the loss of ftsH function results in reduced cytoplasmic retention of the alkaline phosphatase moiety that was attached to cytoplasmic regions of membrane proteins (the Std phenotype) and also in translocation retardation of some exported proteins. We

Rhomboids are a family of serine proteases belonging to intramembrane cleaving proteases, which are supposed to catalyse proteolysis of a substrate protein within the membrane. It remains unclear whether substrates of the rhomboid proteases have a common sequence feature that allows specific cleavage by rhomboids. We showed previously that GlpG, the Escherichia coli rhomboid, can cleave a type I model membrane protein Bla-LY2-MBP having the second transmembrane region of lactose permease (LY2) a

Alkaline phosphatase (PhoA), localized in the periplasmic space of Escherichia coli, is a homodimeric metalloprotein containing two intramolecular disulfide bonds. We attempted to clarify the folding-assembly pathways of this enzyme by allowing in vitro-synthesized PhoA polypeptide to fold into active enzyme and by examining the occurrence of similar pathways in vivo by pulse-chase experiments. PhoA (lacking the signal sequence) that was synthesized in a coupled transcription-translation system

The purpose of the present study is to clarify the effects of hypoxia on the activity of the dopaminergic neurons in the brain and its mechanism of action. For this purpose, the effects of hypoxia on the extracellular levels of 3,4-dihydroxyphenylethylamine (dopamine) were examined in the rat striatum using in vivo brain microdialysis in the presence or absence of pretreatment with either tetrodotoxin (a blocker of voltage-dependent sodium channels) or nomifensine (a blocker of dopamine reuptake