Mechanism and specificity of the human paracaspase MALT1.
The paracaspase domain of MALT1 (mucosa-associated lymphoid tissue lymphoma translocation protein 1) is a component of a gene translocation fused to the N-terminal domains of the cellular inhibitor of apoptosis protein 2. The paracaspase itself, commonly known as MALT1, participates in the NF-κB (nuclear factor κB) pathway, probably by driving survival signals downstream of the B-cell antigen receptor through MALT1 proteolytic activity. We have developed methods for the expression and purification of recombinant full-length MALT1 and its constituent catalytic domain alone. Both are activated by dimerization without cleavage, with a similar dimerization barrier to the distantly related cousins, the apical caspases. By using positional-scanning peptidyl substrate libraries we demonstrate that the activity and specificity of full-length MALT1 is recapitulated by the catalytic domain alone, showing a stringent requirement for cleaving after arginine, and with striking peptide length constraints for efficient hydrolysis. Rates of cleavage (kcat/Km values) of optimal peptidyl substrates are in the same order (10(3)-10(4) M(-1)·s(-1)) as for a putative target protein CYLD. Thus MALT1 has many similarities to caspase 8, even cleaving the putative target protein CYLD with comparable efficiencies, but with diametrically opposite primary substrate specificity.
Sanford-Burnham Medical Research Institute, La Jolla, CA 92037, USA.
Janna
J
Hachmann
Scott J
SJ
Snipas
Bram J
BJ
van Raam
Erik M
EM
Cancino
Emily J
EJ
Houlihan
Marcin
M
Poreba
Paulina
P
Kasperkiewicz
Marcin
M
Drag
Guy S
GS
Salvesen
D000595:Amino Acid Sequence; D020169:Caspases / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification; D002846:Chromatography, Affinity; D002951:Citrates / Q000737:chemistry; D004789:Enzyme Activation; D020536:Enzyme Activators / Q000737:chemistry; D004926:Escherichia coli; D057809:HEK293 Cells; D006801:Humans; D007700:Kinetics; D000074884:Mucosa-Associated Lymphoid Tissue Lymphoma Translocation 1 Protein; D009363:Neoplasm Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification; D009842:Oligopeptides / Q000737:chemistry; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D059748:Proteolysis; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification; D000077559:Sodium Citrate; D013379:Substrate Specificity
D002951:Citrates; D020536:Enzyme Activators; D009363:Neoplasm Proteins; D009842:Oligopeptides; D011993:Recombinant Fusion Proteins; D000077559:Sodium Citrate; D020169:Caspases; C400109:MALT1 protein, human; D000074884:Mucosa-Associated Lymphoid Tissue Lymphoma Translocation 1 Protein