Robert G. Brankamp

Importancia para la hirudoterapia

Robert G. Brankamp's published work in the early 1990s, at the Merrell Dow Research Institute (later Marion Merrell Dow and Hoechst Marion Roussel) in Cincinnati, centred on anticoagulant proteins from the South American leech Haementeria ghilianii. He was a co-author of the 1990 report giving the amino acid sequence of ghilanten, an anticoagulant-antimetastatic principle of 119 amino acids with twenty cysteines, a carboxy-terminal heparin-binding consensus and a reactive arginine-34 implicated in the active-site inhibition of trypsin and factor Xa (PMID 2306252). In 1991 he and colleagues purified five sequence-related variants, P1–P5, demonstrating rapid, stoichiometric inhibition of human factor Xa, and showed that a synthetic carboxy-terminal peptide bound heparin at physiological pH and salt concentration and suppressed lung metastases in mice, though to a lesser extent than whole ghilanten (PMID 1772984). His structural work extended to antistasin, another leech-derived factor Xa inhibitor: a 1992 study in FEBS Letters showed that a synthetic alanine-substituted carboxy-terminal fragment, [A103,106,108]antistasin 93–119, bound sulfatide specifically and saturably, and that glycosaminoglycans blocked this binding in a rank order identical to that for whole antistasin, identifying residues 93–119 as a critical domain for glycosaminoglycan and sulfated-glycolipid binding (PMID 1733769). In 1995 he reported recombinant production of ghilanten, assembling a synthetic gene from eight double-stranded oligonucleotide fragments and expressing it in Pichia pastoris under the methanol-inducible AOX1 promoter; the protease-deficient strain SMD1168 secreted about twofold more product than KM71, and the protein was recovered in a single heparin-Sepharose affinity step (PMID 8746634).