By utilizing two different approaches, we have isolated several proteins which may serve as LPS binding proteins on human monocytes. One method used was to add LPS to 35-S-methionine labeled monocytes, lyse the cells and then, using an anti-LPS antibody, immunoprecipitate the LPS along with any bound monocyte proteins. We have found two proteins, one greater than 200 kd and one approximately a 100 kd in site. The greater than 200 kd protein has been identified by Western blot analysis as CD45. The 110 kd protein has not yet been identified, but we have found that its expression is enhanced by pretreatment with IFN-gamma. The second approach used involved covalently linking LPS to CNBr-activated sepharose beads. The beads are then added to a lysate of 35-S-methionine labeled monocytes and washed several times to remove proteins which do not bind the LPS. Using this technique, we have found two additional proteins which are approximately 70 and 90 kd. Work is currently in progress to identify all these proteins and to verify their role in the LPS induced activation of human monocytes.
