A Diversified Family of 12‐kDa Proteins with a High Amino Acid Sequence Similarity to Macrophage Migration‐Inhibitory Factor (MIF)

Abstract

Two isoforms of a bovine-brain-derived 12-kDa protein (designated p12" and ~1 2~) whose Ntermini have a high amino acid sequence similarity with the glycosylation-inhibiting factor (GIF) and macrophage migration-inhibitory factor (MIF) were purified to homogeneity. The complete amino acid sequence of bovine p12" (PI 9.5) was determined by Edman degradation of the intact molecule and overlapping fragments generated by proteolytic cleavage. The pl2" isoform has nine and ten conservative substitutions versus human GIF (hGIF) and human MIF (hMIF), respectively. Molecular filtration revealed that both isoforms of pl2 exist as monomers in aqueous solution. Circular dichroism spectra indicate that both isoforms of p12 consist of 39% 3% a helix, 23 5 3% p structure and 15 % 3% p turns. Although the N-terminal parts of p12" and ~1 2~ have weak amino acid sequence similarity with that of glutathione S-transferase (GST) neither isoform of p12 was bound to a GST-affinity gel nor had GST activity. Despite a high amino acid sequence similarity with human MIF neither of the p12 isoforms inhibited migration of the mouse monocyte-macrophage cells P338D,.

Crystal structure at 2.6-A resolution of human macrophage migration inhibitory factor.

“…MIF forms a trimer with dimensions of 35 A x 50 A x 50 A. The trimeric arrangement of MIF is consistent with the prediction from a preliminary crystallographic investigation (44) but differs from other reports on the subunit structure of MIF based on gel-filtration or analytical ultracentrifugation experiments that indicate MIF is either a monomer or dimer (3,9,20). The MIF trimer is an a/13 structure with six a-helices surrounding three 13-sheets that completely wrap around to form a barrel with open ends forming a solvent channel (Fig.…”

“…Macrophage migration inhibitory factor (MIF) is a protein of 115 amino acids that is expressed in a wide variety of tissues including the cells of the immune system, anterior pituitary, liver, developing eye lens, and brain (1)(2)(3)(4). MIF is considered to be the first lymphokine discovered and was first described over 30 years ago as a soluble factor produced by activated T cells that inhibits the migration of macrophages (5,6).…”

MIF intersubunit disulfide mutant antagonist supports activation of CD74 by endogenous MIF trimer at physiologic concentrations

“…Previous studies indicate WT MIF is present as a monomer, dimer, or trimer, but most of these experiments were performed at concentrations much higher than the low nanomolar concentrations used physiologically for MIF activation of CD74 (13)(14)(15)(16)(17)(18)(19)(20). Mutagenesis at the subunit interface has not been successful at producing a soluble monomer to test for receptor binding and biological activity (20).…”

“…Although the crystal structure of MIF indicates that it is a trimer, other methods have been used to identify MIF monomers or dimers, in addition to the trimer (13)(14)(15)(16)(17)(18)(19)(20)(21)(22). Sedimentation velocity and equilibrium ultracentrifugation data found MIF to be 97-98% trimeric species with a small amount of a large aggregate and an "incompetent monomer" not in equilibrium with the trimer (23).…”

In Vivo Blockade of Macrophage Migration Inhibitory Factor Ameliorates Acute Experimental Autoimmune Encephalomyelitis by Impairing the Homing of Encephalitogenic T Cells to the Central Nervous System

“…Moreover, elevated levels of MIF were reported in the cerebrospinal fluid of patients with MS (17). MIF is constitutively expressed in the CNS, and it is up-regulated in response to inflammatory and infectious stimuli (18,19). However, very little is known about the role of MIF in the regulation of T cell responses in the CNS, and no direct evidence has been provided for its involvement in the pathogenesis of MS/experimental autoimmune encephalomyelitis (EAE).…”