The F. necrophorum gene expressing leukotoxin was sequenced and cloned.
The leukotoxin open reading frame (lktA) is part of a multi-gene operon
containing 9,726 bp, and encoding a protein containing 3,241 amino acids
with an overall molecular weight of 335,956 daltons. The protein encoded
by the gene was truncated into five polypeptides having overlapping
regions by truncating the full length gene into five different sections
and amplifying, expressing, and recovering the protein encoded by each of
these sections. Additionally, a region upstream of the gene was sequenced
and the polypeptide encoded by that nucleotide sequence was purified and
isolated. These polypeptides along with the full length protein are then
tested to determine their immunogenicity and protective immunity in
comparison to the efficacy of immunization conferred by inactivated
native leukotoxin in F. necrophorum culture supernatant.