Isolation of mammalian calelectrins: a new class of ubiquitous calcium(2+)-regulated proteins

Abstract

In a new approach to isolating proteins which participate in the Ca2+-dependent regulation of membrane traffic in animal cells, two new Ca2+-binding proteins (Mr 67 000 and 32 500) have been identified in and purified from bovine liver, brain, and adrenal medulla. These proteins specifically and reversibly bind to chromaffin granule membranes at low Ca2+ concentrations (half-maximal binding at 5.5 microM Ca2+) and greatly potentiate the Ca2+-induced aggregation of these membranes at higher concentrations (above 10 microM). In the presence of ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetate, the isolated proteins have Stokes radii of 3.40 nm (Mr 67 000) and 2.53 nm (Mr 32 500) as estimated by gel filtration and therefore occur as monomers. They are slightly acidic proteins with pI's of 5.85 and 5.60. In bovine tissues, both proteins and a third protein of Mr 35 000 cross-react immunologically with each other and with Torpedo calelectrin (Mr 34 000) and are therefore identified as mammalian calelectrins. In all tissues of Torpedo marmorata tested, only a single molecular mass form of calelectrin exists, whereas multiple forms of calelectrin exist in mammalian tissues, indicating gene duplication during evolution. We suggest that the evolutionary conservation and diversification, the high tissue concentrations, and the Ca2+-specific interactions of the calelectrins make them candidates for Ca2+-dependent regulators of membrane events in animal cells.

Characterization and ultrastructural localization of annexin VI from mitochondria

“…Mitochondrial annexin VI from bovine liver has the same electrophoretic migration as the form exhibiting the hydrophobic hexapeptide VAAEIL [21]. Its amino acid composition is very similar to those of previously described bovine [18,22,23,24] or human annexins VI [25] and its N-terminus is blocked. Proteolysis ofmitochondrial annexin VI gives rise to peptides identical to those of bovine annexins VI from other sources, as determined by sequence determination [18,25,26].…”

“…1, M), only one band could be observed at the level of the upper band of the doublet. Table 1 Comparison of the amino acid composition (mol %) of the 67 kDa protein isolated from bovine liver mitochondria with those of annexins VI isolated from Bovine liver [22,23], bovine brain [24], or bovine aorta [18], and with the amino acid composition calculated from the sequence of the human annexin VI [25]. …”

Isolation and characterization of two novel calcium‐dependent phospholipid‐binding proteins from bovine lung

“…Recently, a new family of calcium-binding proteins has been identified in a wide range of tissues, and termed the annexins [2,3]. The members of this group so far identified include calelectrin from the electric ray Torpedo marmorata [4,5] and the mammalian proteins calpactin I (a substrate of the tyrosine kinase pp60"' [6,7], lipocortin I (a substrate of the EGF receptor tyrosine kinase) [7,8], endonexin [9,10], protein II [11,12], endonexin II [13] and p70 [9,11,14]. These proteins share the property of binding to acidic phospholipids in the presence of micromolar calcium concentrations [5,9,10,13, 15-181.…”

Purification and characterization of phospholipase A2 inhibitory proteins from pig thyroid gland

“…This protein belongs to a group of calcium-precipitable proteins described previously in many tissues and cells [2,3,5-g]. Biochemical studies (molecular relative mass, isoelectric point and amino acid composition) show that the 32 kDa thyroid protein shares common properties with endonexin [20,37]. Nevertheless we could demonstrate that this protein was recognized by a polyclonal antibody raised against a lipocortin indicating that this 32 kDa thyroid protein belongs to the lipocortin family.…”