Localization and targeting of the Saccharomyces cerevisiae Kre2p/Mnt1p alpha 1,2-mannosyltransferase to a medial-Golgi compartment

Kre2p antibodies were raised in rabbits against a BSA-coupled synthetic peptide corresponding to the last 10 amino acid residues of the protein (NH2-KPKNWKKFRE-COOH; obtained from the Sheldon Biotechnology Centre, McGill University, Montr6al, Qu6bec, Canada). Initially, rabbits were injected with 500 ~g of conjugated peptide in Freund's complete adjuvant, followed by three subsequent injections with equivalent amounts of peptide in Freund's incomplete adjuvant at 3-4-wk intervals. The conjugated Kre2p peptide was coupled to cyanogen bromide-activated Sepharose CL-6B (Pharmacia LKB Biotechnology, Inc., Montr6al, Canada) and used in a column to affinity purify the antiserum as described by Raymond et aL (1990) .