Analysis of a novel gene and beta-galactosidase isozyme from a psychrotrophic Arthrobacter isolate

We have characterized a new psychrotrophic Arthrobacter isolate which produces beta-galactosidase isozymes. When DNA from this isolate was transformed into an Escherichia coli host, we obtained three different fragments, designated 12, 14, and 15, each encoding a different beta-galactosidase isozyme. The beta-galactosidase produced from fragment 12 was of special interest because the protein subunit was smaller (about 71 versus 116 kDa) than those typically encoded by the lacZ family. The isozyme encoded by fragment 12 was purified, and its activity and thermostability were examined. Although the enzyme is highly specific towards beta-D-galactoside substrates, its levels in the isolate do not increase in cells grown with lactose. Nucleotide sequence determination showed that the gene encoding isozyme 12 is not similar to the other members of the lacZ family but has regions similar to beta-galactosidase isozymes from Bacillus stearothermophilus and B. circulans. Addition of the isozyme 12 sequence to the database made it possible to examine these enzymes as possible members of a new, separate family. Our analysis of this new family showed some conserved amino acids corresponding to the lacZ acid-base catalytic region but no homology with the nucleophilic region. On the basis of these comparisons, we designated this a new lacG family.