Evidence for In Vivo Synthesis of Thiamin Triphosphate by Cytosolic Adenylate Kinase in Chicken Skeletal Muscle

Abstract

We showed previously that cytosolic adenylate kinase (AK1) purified from pig skeletal muscle catalyzes in vitro formation of thiamin triphosphate (TTP) from thiamin diphos-phate (TDP) and ADP in addition to ATP formation from ADP [Shikata, H. et al. (1989) Biochem. Int. 18, 933-942]. To obtain evidence for in vivo synthesis of TTP by AK1, changes in TTP content and AK1 activity were determined in chicken skeletal muscle during development after hatching. Thiamin phosphate metabolism in chicken skeletal muscle was also studied. i) An extremely high TTP content, 81% of total thiamin (thiamin plus thiamin phosphates), was detected in the white (fast-twitch) muscle of adult normal chicken (5th to 9th month) compared with a relatively high TTP content of 31% in the red (slow-tonic) muscle. Since approximately equivalent amounts of total thiamin were present in the two types of muscle, the ratio of TTP to TDP was high (5.0) in the white muscle and low (0.41) in the red muscle. ii) Rabbit anti-chicken AK1 antiserum against the purified chicken cytosolic AK1 preparation was obtained. Both AK1 activity and TTP- synthesizing activity in crude cytosol fraction of adult chicken white muscle were inhibited in parallel by the antiserum. iii) In the white muscle of normal chicken, the TTP content and AK1 activity responsible for forming either ATP or TTP were increased in a parallel manner up to day 16 after hatching, after which both remained constant. In the red muscle, on the other hand, both the TTP content and the AK1 activity were low in comparison with those in the white muscle, and were almost constant after hatching. These results suggest in vivo synthesis of TTP by AK1. iv) The TTP/TDP ratio on day 2 after hatching was nearly the same (0.19-0.34) in both types of muscle, and then increased to 3.7 in the white muscle on day 16 after hatching, but remained almost constant in the red muscle. These results suggest that in the chicken, TTP metabolism in the white muscle is initially the same as that in the red muscle just after hatching, and then develops into the white muscle type in the following 2 weeks.