Gonciarz-Swiatek Malgorzata, Rechsteiner Martin
Department of Biochemistry, University of Utah, School of Medicine, 15 North Medical Drive East RM 4100, Salt Lake City, UT 84112-5650, USA.
Mol Immunol. 2006 May;43(12):1993-2001. doi: 10.1016/j.molimm.2005.11.012. Epub 2006 Jan 19.
Previously we proposed that stretches of alternating Lys(K) and Glu(E) in polypeptides promote the expression of nearby sequences on Class I molecules [Realini, C., Rogers, S.W., Rechsteiner, M., 1994. KEKE motifs. Proposed roles in protein-protein association and presentation of peptides by MHC class I receptors. FEBS Lett. 348, 109-113]. As a test of the KEKE hypothesis we have employed osmotic lysis of pinosomes and transfection to introduce or express various ubiquitin peptide fusion proteins in the cytosol of wild type and PA28alphabetagamma- mouse embryo fibroblasts. KEKE or non-KEKE motifs were placed between ubiquitin and the OVA epitope SIINFEKL that was at or near the C-termini of the various fusion proteins. Measurements of surface Kb-SIINFEKL complexes using the monoclonal antibody 25.-D1.16 allowed us to assess the effects of upstream KEKE motifs and PA28 status on SIINFEKL surface presentation. KEKE motifs did not enhance presentation of the OVA epitope. However, our studies did confirm that PA28alphabeta is needed for efficient SIINFEKL surface expression when hsp90 is inhibited.
此前我们提出，多肽中交替出现的赖氨酸（K）和谷氨酸（E）片段可促进I类分子上附近序列的表达[雷亚利尼，C.，罗杰斯，S.W.，雷施泰纳，M.，1994年。KEKE基序。在蛋白质-蛋白质结合以及MHC I类受体呈递肽段中的假定作用。《欧洲生物化学学会联合会快报》348，109 - 113]。作为对KEKE假说的一项检验，我们利用吞噬体的渗透裂解和转染，在野生型和PA28αβγ-小鼠胚胎成纤维细胞的胞质溶胶中引入或表达各种泛素肽融合蛋白。KEKE或非KEKE基序被置于泛素与各种融合蛋白C末端处或附近的OVA表位SIINFEKL之间。使用单克隆抗体25.-D1.16对表面Kb - SIINFEKL复合物进行测量，使我们能够评估上游KEKE基序和PA28状态对SIINFEKL表面呈递的影响。KEKE基序并未增强OVA表位的呈递。然而，我们的研究确实证实，当hsp90受到抑制时，PA28αβ对于有效的SIINFEKL表面表达是必需的。
