A stream of genetic and biochemical information available for the biosynthesis of aminocyclitols over the past few years has provided the foundation to study the modes of formation of this clinically important class of natural products. In addition to work on the identification and functional analysis of aminocyclitol biosynthetic gene clusters, a contingent of recent studies has focused on the detailed analysis of unique enzymatic and catalytic mechanisms inherent to these pathways. The results provide invaluable insights into the biochemical and molecular aspects of aminocyclitol biosynthesis and have revealed diverse and unique features of the pathways.