Endoinulinase from Arthrobacter sp. S37 (EnIA), a member of the glycoside hydrolase family 32, is unique in that, unlike other members of the family, it contains a 250-residue N-terminal domain including a "laminin-G like jelly-roll" fold. This unique N-terminal domain is here suggested to be involved in dimerization and catalysis. The essentially inactive nature of enzymes produced by N-terminal truncation (Delta15, Delta45, Delta70, and Delta250) supported the pivotal role of this unique domain in catalysis and the need for its structural integrity. Significant reductions in the enzyme efficiency (kcat/Km) were observed when mutations were introduced at highly conserved tryptophan residues (Trp75 and Trp141) in the laminin-G like jelly-roll fold, implying their involvement in catalysis. Results from size-exclusion chromatography of the native and chimeric enzymes in the presence and absence of the domain suggested that the N-terminal domain could mediate dimerization.