Loya S, Rudi A, Kashman Y, Hizi A
Department of Cell Biology and Histology, Sackler School of Medicine, Tel Aviv University, Tel Aviv 69978, Israel.
Biochem J. 1997 Jun 15;324 ( Pt 3)(Pt 3):721-7. doi: 10.1042/bj3240721.
A natural compound from the Red Sea sponge Ircinia sp., 2-hexaprenylhydroquinone (HPH), has been shown to be a general inhibitor of retroviral reverse transcriptases (from HIV-1, HIV-2 and murine leukaemia virus) as well as of cellular DNA polymerases (Escherichia coli DNA polymerase I, and DNA polymerases alpha and beta). The pattern of inhibition was found to be similar for all DNA polymerases tested. Thus the mode of inhibition was studied in detail for HIV-1 reverse transcriptase. HPH is a non-competitive inhibitor and binds the enzyme irreversibly with high affinity (Ki=0. 62 microM). The polar hydroxy groups have been shown to be of key importance. A methylated derivative, mHPH, which is devoid of these polar moieties, showed a significantly decreased capacity to inhibit all DNA polymerases tested. Like the natural product, mHPH binds the enzyme independently at an allosteric site, but with reduced affinity (Ki=7.4 microM). We show that HPH does not interfere with the first step of the polymerization process, i.e. the physical formation of the reverse-transcriptase-DNA complex. Consequently, we suggest that the natural inhibitor interferes with the subsequent steps of the overall reaction. Since HPH seems not to affect the affinity of dNTP for the enzyme (the Km is unchanged under conditions where the HPH concentration is increased), we speculate that its inhibitory capacity is derived from its effect on the nucleotidyl-transfer catalytic reaction. We suggest that such a mechanism of inhibition is typical of an inhibitor whose mode of inhibition should be common to all RNA- and DNA-directed polymerases.
一种从红海海绵Ircinia sp.中提取的天然化合物2-己异戊二烯基对苯二酚（HPH），已被证明是逆转录病毒逆转录酶（来自HIV-1、HIV-2和鼠白血病病毒）以及细胞DNA聚合酶（大肠杆菌DNA聚合酶I、DNA聚合酶α和β）的通用抑制剂。研究发现，所有测试的DNA聚合酶的抑制模式相似。因此，对HIV-1逆转录酶的抑制模式进行了详细研究。HPH是一种非竞争性抑制剂，以高亲和力（Ki = 0.62 microM）不可逆地结合该酶。已证明极性羟基至关重要。一种不含这些极性部分的甲基化衍生物mHPH，对所有测试的DNA聚合酶的抑制能力显著降低。与天然产物一样，mHPH在变构位点独立结合该酶，但亲和力降低（Ki = 7.4 microM）。我们表明，HPH不会干扰聚合过程的第一步，即逆转录酶-DNA复合物的物理形成。因此，我们认为这种天然抑制剂会干扰整个反应的后续步骤。由于HPH似乎不影响dNTP对该酶的亲和力（在HPH浓度增加的条件下Km不变），我们推测其抑制能力源于其对核苷酸转移催化反应的影响。我们认为这种抑制机制是一种抑制剂的典型机制，其抑制模式应该对所有RNA和DNA指导的聚合酶都通用。
