DHFR is the most intensively studied enzyme in the folate pathway. It is an essential and ubiquitous enzyme. It generates Tetra Hydro Folate (THF), various cofactors which are involved in the transfer reactions of one carbon unit used in the biosynthesis of nucleic and amino acids, including methylation of dUMP to dTMP. DHFR inhibitors act by halting synthesis of DNA, RNA, and proteins, thereby arresting cell growth. DHFR is an important target for drug development against cancer and a variety of infectious diseases caused by bacteria, protozoa, and fungi. DHFR from Pneumocystis carinii has received significant attention because this pathogen is often associated with AIDS and other immune deficiencies. Molecular modeling techniques are now widely used in medicinal chemistry for design and optimization of inhibitors for a specific target. A pharmacophore model was generated using reported Pc DHFR inhibitors in PHASE module of Schrodinger suite, the pharmacophore model was able to accurately predict Pc DHFR activity. Further the molecules were subjected to molecular docking to understand the binding mode, the docking results also provide additional confidence in the proposed Pharmacophore model. Results suggested that the proposed 3D QSAR model and docking analysis can be useful to rationally design new inhibitors. A combined pharmacophore based and docking based virtual screening was performed to obtain a possible lead molecule for further optimization.