Accession Number : ADA290118

Title :   Cloning and Functional Analysis of Saxiphilin, a Saxitoxin-Binding Protein from the Bullfrog.

Descriptive Note : Midterm rept.,

Corporate Author : YALE UNIV NEW HAVEN CT

Personal Author(s) : Moczydlowski, Edward G.

PDF Url : ADA290118

Report Date : 01 OCT 1994

Pagination or Media Count : 29

Abstract : Saxitoxin (STX), a potent neurotoxin, is the causative agent of paralytic shellfish poisoning in humans. The North American bullfrog, Rana Icatesbeiana, contains a plasma protein called saxiphilin that binds STX with high affinity and specificity. Inasmuch as saxiphilin may be useful as a detection re-agent or an antidote for STX, we have investigated the biochemical properties of saxiphilin and cloned cDNA encoding this protein. Native saxiphilin is a polypeptide of 826 amino acid residues (Mr = 90,818) that contains one binding site for 3HjSTx per inolecule with an equilibrium dissociation constant of KDN 0.2 nM. The amino acid sequence of saxiphilin, deduced from cDNA isolated from bullfrog liver, exhibits substantial homology to members of the transferrin family of Fe3+-binding proteins. However, biochemical and immunochemical analyses confirm that saxiphilin is a unique protein that is not derived from bullfrog serum transferrin. Also, saxiphilin does not bind Fe3+ which implies that saxiphilin is probably not involved in iron metabolism. The mechanism of 3Hstx binding to saxiphilin, including the pH-dependence and temperature-dependence, has been characterized in detail. Recombinant saxiphilin has been expressed in insect cells using a baculovirus vector and the STX-binding site has been localized to the C-lobe domain of the protein.

Descriptors :   *TOXINS AND ANTITOXINS, DETECTION, HUMANS, CELLS, BIOCHEMISTRY, PROTEINS, SITES, EQUILIBRIUM(GENERAL), METABOLISM, DEOXYRIBONUCLEIC ACIDS, SEQUENCES, IRON, FUNCTIONAL ANALYSIS, ANTIDOTES, POISONING, CLONES, GENETIC ENGINEERING, VIRUSES, DISSOCIATION, POTENCY, AMINO ACIDS, CNS DEPRESSANTS, BLOOD PLASMA, INSECTS, BLOOD PROTEINS, IMMUNOCHEMISTRY, PARALYSIS, SHELLFISH, MUSCLE FIBERS.

Subject Categories : Toxicology
      Biology

Distribution Statement : APPROVED FOR PUBLIC RELEASE