A full-length cDNA encoding type II arginase was isolated from a human kidney cDNA library and its sequence compared to those of vertebrate type I arginases as well as to arginases of bacteria, fungi and plants. The predicted sequence of human type II arginase is 58% identical to the sequence of human type I arginase but is 71% identical to the sequence of Xenopus type II arginase, suggesting that duplication of the arginase gene occurred before mammals and amphibians diverged. Seven residues known to be essential for activity were found to be conserved in all arginases. Type II arginase mRNA was detected in virtually all human and mouse RNA samples tested whereas type I arginase mRNA was found only in liver. At least five mRNA species hybridizing to type II arginase cDNA were found in the human RNA samples whereas only a single type II arginase mRNA species was found in the mouse. This raises the possibility that the multiple type II arginase mRNAs in humans arise from differential RNA processing or usage of alternative promoters.