The penicillin-binding proteins are the enzyme catalysts of the critical transpeptidation crosslinking polymerization reaction of bacterial peptidoglycan synthesis and the molecular targets of the penicillin antibiotics. Here, we report a combined crystallographic, small-angle X-ray scattering (SAXS) in-solution structure, computational and biophysical analysis of PBP1 of Staphylococcus aureus (saPBP1), providing mechanistic clues about its function and regulation during cell division. The structure reveals the pedestal domain, the transpeptidase domain, and most of the linker connecting to the “penicillin-binding protein and serine/threonine kinase associated” (PASTA) domains, but not its two PASTA domains, despite their presence in the construct. To address this absence, the structure of the PASTA domains was determined at 1.5 Å resolution. Extensive molecular-dynamics simulations interpret the PASTA domains of saPBP1 as conformationally mobile and separated from the transpeptidase domain. This conclusion was confirmed by SAXS experiments on the full-length protein in solution. A series of crystallographic complexes with β-lactam antibiotics (as inhibitors) and penta-Gly (as a substrate mimetic) allowed the molecular characterization of both inhibition by antibiotics and binding for the donor and acceptor peptidoglycan strands. Mass-spectrometry experiments with synthetic peptidoglycan fragments revealed binding by PASTA domains in coordination with the remaining domains. The observed mobility of the PASTA domain in saPBP1 could play a crucial role for in vivo interaction with its glycosyltransferase partner in the membrane or with other components of the divisome machinery, as well as for coordination of transpeptidation and polymerization processes in the bacterial divisome.

青霉素结合蛋白是细菌肽聚糖合成关键转肽交联聚合反应的酶催化剂，是青霉素抗生素的分子靶点。这里，我们报告一个合并晶体，小角度X射线散射（SAXS）中的溶液结构，计算和生物物理分析 的PBP1金黄色葡萄球菌（SA  PBP1)，提供有关其在细胞分裂过程中的功能和调节的机制线索。该结构揭示了基座结构域、转肽酶结构域和大部分连接到“青霉素结合蛋白和丝氨酸/苏氨酸激酶相关”（PASTA）结构域的接头，但没有显示其两个 PASTA 结构域，尽管它们存在于构建体中。为了解决这种缺失，PASTA 域的结构 以 1.5 Å 分辨率确定。广泛的分子动力学模拟将sa PBP1的 PASTA 域解释为构象可移动并与转肽酶域分离。该结论通过对溶液中全长蛋白质的 SAXS 实验得到证实。    一系列与 β-内酰胺抗生素（作为抑制剂）和五甘氨酸（作为底物模拟物）的晶体复合物允许对抗生素的抑制作用和供体和受体肽聚糖链的结合进行分子表征。使用合成肽聚糖片段的质谱实验揭示了 PASTA 域与其余域的结合。观察到的 PASTA 结构域在sa PBP1 中的移动性对于与膜中的糖基转移酶伙伴或与二分体机械的其他成分的体内相互作用以及细菌二分体中转肽和聚合过程的协调可能起着至关重要的作用。