Biophysical characterization of macrophage migration enhancement factor (MEF).
Macrophage migration enhancement factor (MEF), a lymphokine produced in the spleen by suppressor-like lymphoid cells, may be an important immunoregulatory molecule of macrophage function. MEF appears to be a potent positive chemokinetic factor and is unusual in that it lacks chemotactic activity. To aid in the development of a purification scheme for MEF we have employed biophysical characterization techniques to define its physical properties. Using the technique of velocity sedimentation in isokinetic sucrose gradients, the S20w for MEF was determined to be 2.25. The Stoke's radius for MEF was determined by Sephadex G-100 gel filtration to be 28.9 A. From these measurements the D20w was calculated to be 7.55 x 10(-7) cm2/sec, the mol. wt was calculated to be 28,000, the frictional ratio (f/f0) was calculated to be 1.45, the axial ratio was calculated to be 1:8, and the dimensions of the molecule were estimated to be 20 x 160 A. Using the technique of isoelectric focusing in liquid density gradients, the isoelectric point for MEF was estimated to be 8.8. We have also determined by enzyme treatment that MEF is resistant to DNase and RNase and susceptible to proteinase K and L-fucosidase. In addition, MEF partitioned to the aqueous phase during methanol-chloroform extraction procedures. MEF was inactivated at pH 12; at 100 degrees C MEF was stable for 10 min but was inactive after 1 hr. Collectively, these data will facilitate the development of a purification scheme for MEF which will ultimately permit the analysis of the molecule and its function.
Department of Microbiology and Immunology, Bowman Gray School of Medicine, Wake Forest University Medical Center, Winston-Salem, NC 27103.
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Gordon
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Myrvik
D000818:Animals; D002499:Centrifugation, Density Gradient; D002630:Chemotactic Factors / Q000302:isolation & purification; D002633:Chemotaxis; D002850:Chromatography, Gel; D004355:Drug Stability; D004798:Enzymes / Q000494:pharmacology; D005590:Fractional Precipitation; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D008264:Macrophages / Q000276:immunology*; D008970:Molecular Weight; D011817:Rabbits; D013696:Temperature
D002630:Chemotactic Factors; D004798:Enzymes; C053609:macrophage migration enhancement factor