Ramasamy M, Alpers D H, Tiruppathi C, Seetharam B
Department of Medicine, School of Medicine, Washington University, St. Louis, Missouri 63110.
Am J Physiol. 1989 Nov;257(5 Pt 1):G791-7. doi: 10.1152/ajpgi.1989.257.5.G791.
To ascertain the mechanism of release of cobalamin (Cbl) from intrinsic factor (IF) and subsequent formation of transcobalamin II (TC-II)-Cbl complex, we studied the intracellular distribution of 57Co-labeled Cbl after its uptake in suckling and adult rats. The amount of Cbl bound to IF, to the IF-Cbl receptor via IF, and to TC-II was determined by immunoprecipitation with monospecific antisera raised to these proteins. IF-Cbl receptor activity was found to be very low in suckling rats up to 12 days after birth. Oral administration of leupeptin in amounts known to alter protein turnover had no effect on the release of Cbl from IF nor did it inhibit the formation of the TC-II-Cbl complex in either adult or suckling animals. However, oral administration of chloroquine resulted in a transient increase in the intestinal concentration of Cbl in both adult and suckling rats and in total inhibition of Cbl released from IF in adults rats. Chloroquine prevented completely the transfer of Cbl to TC-II in adult rats and inhibited the transfer by 50% in suckling rats. These data demonstrate that in adult mucosa utilizing receptor-mediated endocytosis, Cbl is transferred from IF to TC-II. This transfer does not require the IF-Cbl receptor, as it occurs in suckling rats. Finally, transfer of Cbl to TC-II is decreased by a drug that alters vesicular pH. Because Cbl can be released at acid pH from IF, it is proposed that release of Cbl from IF and its transfer to TC-II occurs in an acidic vesicle.
为了确定钴胺素（Cbl）从内因子（IF）释放以及随后转钴胺素II（TC-II）-Cbl复合物形成的机制，我们研究了57Co标记的Cbl在乳鼠和成年大鼠体内摄取后的细胞内分布。通过用针对这些蛋白质产生的单特异性抗血清进行免疫沉淀，测定了与IF结合、通过IF与IF-Cbl受体结合以及与TC-II结合的Cbl量。发现出生后12天内的乳鼠中IF-Cbl受体活性非常低。口服已知会改变蛋白质周转的亮抑酶肽，对Cbl从IF的释放没有影响，也不抑制成年或乳鼠中TC-II-Cbl复合物的形成。然而，口服氯喹导致成年和乳鼠肠道中Cbl浓度短暂升高，并完全抑制成年大鼠中从IF释放的Cbl。氯喹完全阻止了成年大鼠中Cbl向TC-II的转移，并在乳鼠中抑制了50%的转移。这些数据表明，在利用受体介导的内吞作用的成年黏膜中，Cbl从IF转移到TC-II。这种转移不需要IF-Cbl受体，因为它在乳鼠中发生。最后，一种改变囊泡pH值的药物会降低Cbl向TC-II的转移。由于Cbl在酸性pH值下可从IF释放，因此提出Cbl从IF的释放及其向TC-II的转移发生在酸性囊泡中。
