Structural organization of the gene encoding the human lipocalin tear prealbumin and synthesis of the recombinant protein in Escherichia coli

Abstract

The genomic DNA fragment encoding the human lipocalin tear prealbumin (LCN1), a new member of the superfamily of hydrophobic molecule transporters, has been isolated and sequenced. The entire gene is approximately 6.2 kb in size and contains six protein-coding exons and a 3′-nontranslated exon. All exon/intron splice junctions exactly follow the GT/AG rule. The structure of the LCN1 gene is highly similar, in terms of numbers and sizes of exons and in intron phasing, to that of the genes encoding ovine β-lactoglobulin, human placental protein P14, rat α2-urinary globulin, rat prostaglandin D synthase and human α1-microgobulin, thus supporting the close evolutionary relationship of these genes. The 5′-noncoding region of LCN1 contains, besides a TATA and CAAT box, several motifs that resemble regulatory elements of other eukaryotic genes, including potential metal-responsive elements ( MRE) and a cAMP-responsive element ( CRE). As a basis for further investigations concerning the structure-function relationship and to generate a source of recombinant protein for X-ray crystallography studies, LCN1 was produced in Escherichia coli as a fusion with maltose-binding protein.