Dihydrofolate reductase (DHFR) was discovered in the late 1950s by investigators searching for folate-dependent enzymes involved in 1-carbon metabolism, with its already known application as anti-cancer and as antibiotics. This study focuses on structure of Dihydrofolate, its interaction with specific amino acids, mechanism of DHFR catalysis. The reason of study was to focus on compounds which can inhibit DHFR and has applications as antifungal agents, antimalarial agents, ant tuberculosis agents, for Leishmaniasis and Trypanosomiasis treatment. Also, to find out the reason of resistance to antifolates and ways to overcome them, recent drugs under each class of agents and drugs under clinical trials. INTRODUCTION: Sidney Futterman fractionated a chicken liver extract and obtained a preparation that catalyzed the reduction of both folate and DHF to THF in 1944. He also found that Methotrexate and Aminopterin, another anticancer drug, inhibited both of these processes. In 1988 Hitchings, Elion and Black were honoured with Nobel Prize in physiology in medicine for selective binding of Trimethoprim and other pharmaceuticals to target dihydrofolate 1. However molecular nature of selectivity still not understood in spite of fact that DHFR is the most thoroughly and comprehensively studied enzymes most fundamental mechanism of functioning of DHFR and its nature of interactions with the inhibitors need to be understood 1. DHFR is found ubiquitously in all dividing cells of prokaryotes and eukaryotes.