Band 3 is the most abundant integral protein of the red blood cell membrane. It performs two critical biological functions: maintaining ionic homeostasis, by transporting Cl- and HCO3-ions, and providing mechanical stability to the erythroid membrane. Erythroid band 3 (AE1) is one of three anion exchangers that are encoded by separate genes. The AE1 gene is transcribed by two promoters: the upstream promoter produces erythroid band 3, whereas the downstream promoter initiates transcription of the band 3 isoform in kidney. To assess the biological consequences of band 3 deficiency, we have selectively inactivated erythroid but not kidney band 3 by gene targeting in mice. Although no death in utero occurred, the majority of homozygous mice die within two weeks after birth. The erythroid band 3 null mice show retarded growth, spherocytic red blood cell morphology and severe haemolytic anaemia. Remarkably, the band 3-/- red blood cells assembled normal membrane skeleton thus challenging the notion that the presence of band 3 is required for the stable biogenesis of membrane skeleton. The availability of band 3-/- mice offers a unique opportunity to investigate the role of erythroid band 3 in the regulation of membrane-skeletal interactions, anion transport and the invasion and growth of malaria parasite into red blood cells.

A mutant Escherichia coli (Ppcc') which was unable to grow on glucose as a sole carbon source was isolated. This mutant had very low levels of phosphoenolpyruvate carboxylase activity (approximately 5% of the wild type). Goat immunoglobulin G prepared against wild-type phosphoenolypyruvate carboxylase cross-reacted with the Ppccenzyme. The amount of enzyme protein in the mutant cells was similar to that found in wild-type cells, but it had greatly diminished specific activity. The catalytically less active mutant enzyme retained the ability to interact with fructose 1,6-bisphosphate, but did not exhibit stabilization of the tetrameric form by aspartate. The pl of the mutant protein was lower (4.9) than that of the wild-type protein (5.1). After electrophoresis and immunoblotting of the partially purified protein, several immunostaining bands were seen in addition to the main enzyme band. A novel method for showing that these bands represented proteolytic fragments of phosphoenolpyruvate carboxylase was developed.