Phytoplasmas are intracellular bacterial plant pathogens that cause devastating diseases in crops and ornamental plants by the secretion of effector proteins. One of these effector proteins, termed SECRETED ASTER YELLOWS WITCHES’ BROOM PROTEIN 54 (SAP54), leads to the degradation of a specific subset of floral homeotic proteins of the MIKC-type MADS-domain family via the ubiquitin-proteasome pathway. In consequence, the developing flowers show the homeotic transformation of floral organs into vegetative leaf-like structures. The molecular mechanism of SAP54 action involves binding to the keratin-like domain of MIKC-type proteins and to some RAD23 proteins, which translocate ubiquitylated substrates to the proteasome. The structural requirements and specificity of SAP54 function are poorly understood, however. Here, we report, based on biophysical and molecular biological analyses, that SAP54 folds into an α-helical structure. Insertion of helix-breaking mutations disrupts correct folding of SAP54 and compromises SAP54 binding to its target proteins and, concomitantly, its ability to evoke disease phenotypes in vivo. Interestingly, dynamic light scattering data together with electrophoretic mobility shift assays suggest that SAP54 preferentially binds to multimeric complexes of MIKC-type proteins rather than to dimers or monomers of these proteins. Together with data from literature, this finding suggests that MIKC-type proteins and SAP54 constitute multimeric α-helical coiled coils. Our investigations clarify the structure-function relationship of an important phytoplasma effector protein and may thus ultimately help to develop treatments against some devastating plant diseases.

植原体是细胞内细菌植物病原体，其通过分泌效应蛋白而在作物和观赏植物中引起毁灭性疾病。这些效应蛋白之一，被称为“分泌型紫黄色巫婆的布鲁蛋白54（SAP54）”，通过泛素-蛋白酶体途径导致MIKC型MADS结构域家族的花同源蛋白的特定子集降解。结果，正在发育的花朵显示出花器官向植物性叶状结构的同源转化。SAP54作用的分子机制涉及结合MIKC型蛋白的角蛋白样结构域和某些RAD23蛋白，后者将泛素化的底物转位到蛋白酶体上。但是，对SAP54功能的结构要求和特异性了解得很少。在这里，我们报告，根据生物物理和分子生物学分析，SAP54折叠成α螺旋结构。螺旋破坏突变的插入破坏了SAP54的正确折叠，并破坏了SAP54与其靶蛋白的结合，并因此破坏了它在体内引起疾病表型的能力。有趣的是，动态光散射数据与电泳迁移率漂移分析一起表明SAP54优先结合MIKC型蛋白质的多聚体复合物，而不是这些蛋白质的二聚体或单体。连同文献数据一起，该发现表明MIKC型蛋白和SAP54构成了多聚α螺旋卷曲线圈。