The mammalian cytochromes P450 (P450/CYP) from families 1, 2, and 3 play a central role in xenobiotic metabolism. At least 15 different members are expressed in the endoplasmic reticulum (ER) of human liver. Protein-protein interactions involving the different forms of P450 can have dramatic effects on catalytic activities. In order to assess the specificity of these interactions in rat liver microsomes, two P450s (CYP1A2 and CYP3A) were immunoprecipitated (IP), and the proteins co-IP with each form were identified LC-MS profiling. Both P450s showed promiscuous interaction with a variety of P450s and other drug metabolizing enzymes. Gene ontology analysis showed the binding partners of both P450s were representative of catabolic and lipid metabolic biological processes (due in large part to the high number of co-IP P450s) in addition to the responses to drugs and xenobiotic stimuli. Interestingly, the binding partners of CYP1A2 were significantly over-represented with respect to the biological processes of protein folding and the ER-associated unfolded protein response; whereas CYP3A partners were over-represented with respect to sterol and amide biosynthetic processes as well as N-linked protein glycosylation. Comparison to total ER showed that the binding partners of CYP1A2 were associated with higher percentages of proteins related to development, the movement of subcellular components, protein folding, and vesicle targeting, whereas those of P450 3A were associated with higher percentages of proteins related to the movement of subcellular components, and cellular homeostasis. With respect to the percent distribution of molecular functions, CYP1A2 and CYP3A binding partners were similar to the total ER. However, the binding partners of CYP3A were associated with higher percentages of proteins related to transducer and structural molecule activities. Interestingly, the partners of CYP1A2 represented by lower percentages of these two functions. Although there is considerable overlap regarding the processes and functions associated with the binding partners of the two P450s, there appears to be specificity, and perhaps reciprocity, in some cases.