The human APOBEC3G protein is a cytidine deaminase that generates cytidine to deoxy-uridine mutations in single-stranded DNA (ssDNA), and capable of restricting replication of HIV-1 by generating mutations in viral genome. The mechanism by which APOBEC3G specifically deaminates 5\u27-CC motifs has remained elusive since structural studies have been hampered due to apparently weak ssDNA binding of the catalytic domain of APOBEC3G. We overcame the problem by generating a highly active variant with higher ssDNA affinity. Here, we present the crystal structure of this variant complexed with a ssDNA substrate at 1.86 A resolution. This structure reveals atomic-level interactions by which APOBEC3G recognizes a functionally-relevant 5\u27-TCCCA sequence. This complex also reveals a key role of W211 in substrate recognition, implicating a similar recognition in activation-induced cytidine deaminase (AID) with a conserved tryptophan
