The 5α-reduction of testosterone in target tissues is a key step in androgen physiology; however, 5α-reduced C 1 9 steroids are sometimes synthesized in testis via a pathway that does not involve testosterone as an intermediate. We studied the metabolism of 5α-reduced C 2 1 steroids by human cytochrome P450c17 (hCYP17), the enzyme responsible for conversion of C 2 1 steroids to C 1 9 steroids via its 17α-hydroxylase and 17,20-lyase activities. hCYP17 17α-hydroxylates 5α-pregnan-3,20-dione, but little androstanedione is formed by 17,20-lyase activity. hCYP17 also 17α-hydroxylates 5α-pregnan-3α-ol-20-one and the 5α-pregnan-3α,17α-diol-20-one intermediate is rapidly converted to androsterone by 17,20-lyase activity. Furthermore, 5α-pregnan-3α,17α-diol-20-one is a better substrate for the 17,20-lyase reaction than the preferred substrate 17α-hydroxypregnenolone and cytochrome b 5 stimulates androsterone formation only 3-fold. Both 5α-pregnan-3α-ol-20-one and 5α-pregnan-3α,17α-diol-20-one bind to hCYP17 with higher affinity than does progesterone. We conclude that 5α-reduced, 3α-hydroxy-C 2 1 steroids are excellent, high-affinity substrates for hCYP17. The brisk metabolism of 5α-pregnan-3α,17α-diol-20-one to androsterone by CYP17 explains how, when 5α-reductases are present, the testis can produce C 1 9 steroids androsterone and androstanediol from 17α-hydroxyprogesterone without the intermediacy of androstenedione and testosterone.