In the course of the study of the primary structures and molecular mechanis
ms of action of immunologically active compounds of the nervous system we h
ave isolated from the soluble fraction of total bovine brain two heat-stabl
e proteins. The purification procedure was mainly based on DEAE-Servacel io
n-exchange chromatography and reversed-phase HPLC. The proteins were identi
fied by the N-terminal Edman microsequence analysis and database searching
as macrophage migration inhibitory factor (MIF). The N-terminal sequences f
or MIF1 and MIF2 were found to be identical. According to mass spectral ana
lysis, the molecular masses for MIF1 and MIF2 were determined respectively
as 12,369.21 and 12,299.7 Da. In addition, we have also isolated a third pe
ptide having the same N-terminal sequence and Mr 9,496.2 that seems to be a
proteolytic fragment of MIF. Using p-hydroxyphenylpyruvate as a substrate,
we have not revealed tautomerase activity of either MIF1 or MIF2. As both
the immunologic and enzymatic activities were reported to be expressed by t
he oligomeric structure of MIF, we suggest that the present study may give
additional information on MIF in terms of structural properties of this pro
tein. A comparatively simple purification procedure is presented that may b
e widely used for simultaneous isolation in one run of MIF isoforms.