Primary structure and evolution of the ATP-binding domains of the P-type ATPases in Tetrahymena thermophila

Am J Physiol. 1997 Feb;272(2 Pt 1):C715-28. doi: 10.1152/ajpcell.1997.272.2.C715.

Abstract

The P-type ATPases (e.g., Na+-K+-ATPase and Ca2+-ATPase) occur widely in living cells of fungi, Protozoa, plants, and animals. These ion pumps show a high degree of divergence in their primary structures but share a limited number of common amino acid residues for their ATP-catalytic function. Particularly, the amino acid sequences for the phosphorylation site (DKTGTLT) and the binding site for ATP (and its analogs; GDGVND) are conserved throughout evolution. Using two degenerate oligonucleotides corresponding to these regions, we applied a polymerase chain reaction (PCR) technique to the search for P-type ATPase isoforms, which will provide a clue to the evolutionary mechanisms of ion pumps in Tetrahymena thermophila. A total of 12 distinct P-type ATPase genes were identified. Sequence comparisons revealed that seven of them can be compiled into a multigene family, which is similar to animal Na+-K+- and H+-K+-ATPase genes. One of them is close to the sarco(endo)plasmic reticulum Ca2+-ATPase gene, and the other four share a significant homology with the gene encoding Plasmodium ATPase-1 whose function is unknown. A Northern blot analysis and reverse transcriptase-PCR demonstrated that all identified genes are expressed, but the expression levels vary widely under different culture conditions. A Southern blot analysis after pulse-field gel electrophoresis showed that all of these genes exist in T. thermophila macronuclei. The Na+-K+- and H+-K+-ATPase gene family has a high multiplicity (at least 10 different genes detected on genomic Southern blot analysis) and is distributed on four different macronuclear chromosomes. On the basis of a calculation with the amino acid sequences of the cloned cytoplasmic loop region (between the phosphorylation and the gamma-[4-(N-2-chloroethyl-N-methylamino)]-benzylamido ATP sites), the genes with >80% identity form a cognate linkage group within the same macronuclei chromosome, whereas the genes with <70% identity are separated in different chromosomes. The phylogenetic analysis showed that this multigene family is the result of a series of gene duplications.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Adenosine Triphosphatases / chemistry*
  • Adenosine Triphosphatases / genetics
  • Adenosine Triphosphatases / metabolism*
  • Adenosine Triphosphate / metabolism*
  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Chromosomes
  • Evolution, Molecular*
  • Gene Dosage
  • Genes
  • Genetic Linkage
  • Isoenzymes / chemistry*
  • Isoenzymes / genetics
  • Isoenzymes / metabolism*
  • Molecular Sequence Data
  • Tetrahymena thermophila / enzymology*

Substances

  • Isoenzymes
  • Adenosine Triphosphate
  • Adenosine Triphosphatases

Associated data

  • GENBANK/U41357
  • GENBANK/U41358
  • GENBANK/U41359
  • GENBANK/U41360
  • GENBANK/U41361
  • GENBANK/U41362
  • GENBANK/U41363
  • GENBANK/U41364
  • GENBANK/U41365
  • GENBANK/U41366
  • GENBANK/U41367