MOLECULAR-CLONING AND CHARACTERIZATION OF AQPZ, A WATER CHANNEL FROM ESCHERICHIA-COLI

The aquaporin family of molecular water channels is widely expressed t hroughout the plant and animal kingdoms. No bacterial aquaporins are k nown; however, sequence-related bacterial genes have been identified t hat encode glycerol facilitators (glpF), By homology cloning, a novel aquaporin-related DNA (aqpZ) was identified that contained no surface N-glycosylation consensus.The aqpZ RNA was not identified in mammalian mRNA by Northern analysis and exhibited bacterial codon usage prefere nces. Southern analysis failed to demonstrate aqpZ in mammalian genomi c DNA, whereas a strongly reactive DNA was present in chromosomal DNA from Escherichia coli and other bacterial species and did not correspo nd to glpF. The aqpZ DNA isolated from E. coli contained a 693-base pa ir open reading frame encoding a polypeptide 28-38% identical to known aquaporins. When compared with other aquaporins, aqpZ encodes a 10-re sidue insert preceding exofacial loop C, truncated NR(2) and COOH term ini, and no cysteines at known mercury-sensitive sites. Expression of aqpZ cRNA conferred Xenopus oocytes with a 15-fold increase in osmotic water permeability, which was maximal after 5 days of expression, was not inhibited with HgCl2, exhibited a low activation energy (E(a) = 3 .8 kcal/mol), and failed to transport nonionic solutes such as urea an d glycerol. In contrast, oocytes expressing glpF transported glycerol but exhibited limited osmotic water permeability. Phylogenetic compari son of aquaporins and homologs revealed a large separation between aqp Z and glpF, consistent with an ancient gene divergence.