Susceptibility of mitogen-activated protein kinase kinase family members to proteolysis by anthrax lethal factor.
The lethal factor (LF) produced by toxigenic strains of Bacillus anthracis is a Zn(2+)-endopeptidase that cleaves the mitogen-activated protein kinase kinases (MAPKKs) MEK1, MEK2 and MKK3. Using genetic and biochemical approaches, we have extended the study of LF proteolytic specificity to all known MAPKK family members and found that LF also cleaves MKK4, MKK6 and MKK7, but not MEK5. The peptide bonds hydrolysed by LF within all MAPKKs were identified. Cleavage invariably occurs within the N-terminal proline-rich region preceding the kinase domain, thus disrupting a sequence involved in directing specific protein-protein interactions necessary for the assembly of signalling complexes. Alignment of the sequences flanking the site of cleavage reveals the occurrence of some consensus motifs: position P2 and P1' are occupied by hydrophobic residues and at least one basic residue is present between P4 and P7. The implications of these findings for the biochemical activity and functional specificity of LF are discussed.
Vitale
G
G
Centro CNR Biomembrane and Dipartimento di Scienze Biomediche, Università di Padova, Via Trieste 75, 35121 Padova, Italy. gvitale@makek.dstb.uniud.it
Bernardi
L
L
Napolitani
G
G
Mock
M
M
Montecucco
C
C
D000595:Amino Acid Sequence; D000942:Antigens, Bacterial; D001408:Bacillus anthracis / Q000737:chemistry*; D001427:Bacterial Toxins / Q000235:genetics / Q000378:metabolism*; D002384:Catalysis; D006367:HeLa Cells; D006801:Humans; D048371:MAP Kinase Kinase 3; D048670:MAP Kinase Kinase 4; D020929:Mitogen-Activated Protein Kinase Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D005810:Multigene Family; D010802:Phylogeny; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D015398:Signal Transduction; D013379:Substrate Specificity; D020798:Two-Hybrid System Techniques
D000942:Antigens, Bacterial; D001427:Bacterial Toxins; C030325:anthrax toxin; D011505:Protein-Tyrosine Kinases; D048371:MAP Kinase Kinase 3; D048670:MAP Kinase Kinase 4; C482530:MAP2K3 protein, human; C482559:MAP2K4 protein, human; D020929:Mitogen-Activated Protein Kinase Kinases