Thanos C D, Faham S, Goodwill K E, Cascio D, Phillips M, Bowie J U
UCLA-DOE Laboratory of Structural Biology, Department of Chemistry and Biochemistry, UCLA, Los Angeles, California 90095, USA.
J Biol Chem. 1999 Dec 24;274(52):37301-6. doi: 10.1074/jbc.274.52.37301.
The sterile alpha motif (SAM) domain is a protein module found in many diverse signaling proteins. SAM domains in some systems have been shown to self-associate. Previous crystal structures of an EphA4-SAM domain dimer (Stapleton, D., Balan, I., Pawson, T., and Sicheri, F. (1999) Nat. Struct. Biol. 6, 44-49) and a possible EphB2-SAM oligomer (Thanos, C. D., Goodwill, K. E., and Bowie, J. U. (1999) Science 283, 833-836) both revealed large interfaces comprising an exchange of N-terminal peptide arms. Within the arm, a conserved hydrophobic residue (Tyr-8 in the EphB2-SAM structure or Phe-910 in the EphA4-SAM structure) is anchored into a hydrophobic cleft on a neighboring molecule. Here we have solved a new crystal form of the human EphB2-SAM domain that has the same overall SAM domain fold yet has no substantial intermolecular contacts. In the new structure, the N-terminal peptide arm of the EphB2-SAM domain protrudes out from the core of the molecule, leaving both the arm (including Tyr-8) and the hydrophobic cleft solvent-exposed. To verify that Tyr-8 is solvent-exposed in solution, we made a Tyr-8 to Ala-8 mutation and found that the EphB2-SAM domain structure and stability were only slightly altered. These results suggest that Tyr-8 is not part of the hydrophobic core of the EphB2-SAM domain and is conserved for functional reasons. Cystallographic evidence suggests a possible role for the N-terminal arm in oligomerization. In the absence of a direct demonstration of biological relevance, however, the functional role of the N-terminal arm remains an open question.
无菌α基序（SAM）结构域是一种存在于多种不同信号蛋白中的蛋白质模块。在某些系统中，SAM结构域已被证明会发生自我缔合。先前EphA4 - SAM结构域二聚体（Stapleton, D., Balan, I., Pawson, T., and Sicheri, F. (1999) Nat. Struct. Biol. 6, 44 - 49）以及一种可能的EphB2 - SAM寡聚体（Thanos, C. D., Goodwill, K. E., and Bowie, J. U. (1999) Science 283, 833 - 836）的晶体结构均显示出由N端肽臂交换构成的大界面。在该臂内，一个保守的疏水残基（在EphB2 - SAM结构中为Tyr - 8，在EphA4 - SAM结构中为Phe - 910）锚定在相邻分子的疏水裂隙中。在此，我们解析了人EphB2 - SAM结构域的一种新晶体形式，其具有相同的整体SAM结构域折叠，但不存在大量分子间接触。在新结构中，EphB2 - SAM结构域的N端肽臂从分子核心伸出，使得该臂（包括Tyr - 8）和疏水裂隙均暴露于溶剂中。为了验证Tyr - 8在溶液中是否暴露，我们将Tyr - 8突变为Ala - 8，发现EphB2 - SAM结构域的结构和稳定性仅略有改变。这些结果表明，Tyr - 8并非EphB2 - SAM结构域疏水核心的一部分，其保守性是出于功能原因。晶体学证据表明N端臂在寡聚化中可能发挥作用。然而，在缺乏生物相关性直接证据的情况下，N端臂的功能作用仍是一个悬而未决的问题。
