Metal ion content of dolichos biflorus lectin and effect of divalent cations on lectin activity

The metal ion content of the Dolichos biflorus seed lectin has been determined by using atomic absorption spectrophotometry. Ca2+, Mg2+, Mn2+, Zn2+, and Cu2+ were all found in the native lectin and could be removed to different degrees by using a variety of techniques such as high ionic strength, low pH, chelating agent, and combinations of these procedures. By use of affinity electrophoresis, the remaining binding capacity of the lectin could easily be determined, and the D. biflorus lectin showed an absolute requirement for divalent cations to be able to bind N-acetyl-D-galactosamine. The association constant (Ka) of the lectin for N-acetyl-D-galactosamine was determined after remetallization of the lectin with each individual metal ion and a combination of them. The Ka of the lectin for the hapten differed depending on which divalent cation had been used for remetallization. Ca2+ ion alone was equal to a combination of all metal ions in its ability to confer the highest binding activity of the lectin for N-acetyl-D-galactosamine. The other cations, Mg2+, Mn2+, Zn2+, and Cu2+, could not restore the binding activity of the lectin to the same degree as Ca2+ or a combination of all ions. The D. biflorus lectin is, therefore, one of the first lectins that has been shown to require only a single cation to fully retain its binding activity in contrast to most lectins that require a combination of Ca2+, Mn2+, Mg2+, or Zn2+. Furthermore, we have shown that there is a selective process of ion uptake into the lectin since the metal ion ratios in the native lectin compared to the whole seed are quite different.