Molecular cloning and functional characterization of swine sodium dependent phosphate cotransporter type II b (NaPi-IIb) gene.
A sodium-dependent phosphate transporter gene, NaPi-IIb, was isolated from swine small intestine using cDNA library screening method. Sequencing analysis revealed that the NaPi-IIb cDNA sequences was 2,016 bp in length and encoded an open-reading frame consisting of 671 amino acids. The cDNA showed 83.1 % sequences identity to the human NaPi-IIb and 78.7 % sequences identity to the chicken NaPi-IIb. Prediction of membrane spanning domains based on the hydrophilic and hydrophobic properties of the amino acids suggested that a putative protein had nine transmembrane domains, with both the NH(2) and COOH terminal being intracellular. By northern blot, a ~4.2 kb transcript was found to be abundantly expressed in mall intestine, lung, ovary, mammary glands, liver, kidney, salivary glands, placenta and thymus. Microinjection of swine NaPi-IIb cRNA into Xenopus oocytes demonstrated that the NaPi-IIb showed sodium-dependent Pi cotransport activity, and an approximate 31-fold increase of Pi uptake was seen in cRNA injected oocytes. The swine NaPi-IIb transporter expressed in Xenopus oocytes had a Km for Pi of ~79.35 ± 7.2 μM. Furthermore, the pH dependency characterization of swine NaPi-IIb transporter showed activation at extracellular alkaline-pH.
College of Animal Science and Technology, Hunan Agricultural University, Changsha, 410128, Hunan Province, China. zhifeng7809@yahoo.com.cn
Xiang
X
Zhifeng
Fang
F
Rejun
Hu
H
Longchang
Su
S
Wenqing
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D003001:Cloning, Molecular; D020869:Gene Expression Profiling; D006801:Humans; D008969:Molecular Sequence Data; D009865:Oocytes / Q000378:metabolism; D009928:Organ Specificity / Q000235:genetics; D016415:Sequence Alignment; D050614:Sodium-Phosphate Cotransporter Proteins, Type IIb / Q000737:chemistry / Q000235:genetics*; D034421:Sus scrofa / Q000235:genetics*; D014981:Xenopus
D050614:Sodium-Phosphate Cotransporter Proteins, Type IIb