Protein Crystallography Section, Radiation Biology & Health Sciences Division, Bhabha Atomic Research Centre, Trombay, Mumbai, 400085, India; Homi Bhabha National Institute, Anushaktinagar, Mumbai, 400094, India.
Homi Bhabha National Institute, Anushaktinagar, Mumbai, 400094, India; Food Microbiology Group, Food Technology Division, Bhabha Atomic Research Centre, Trombay, Mumbai, 400085, India.
Arch Biochem Biophys. 2023 Mar 1;736:109524. doi: 10.1016/j.abb.2023.109524. Epub 2023 Jan 27.
Alkaline phosphatases (APs), represented by E. coli AP (ECAP), employ an arginine residue to stabilize the phosphoryl group in the active site; whereas, AP from Sphingomonas (SPAP) shows a unique combination of substrate-binding residues; Thr89, Asn110, Lys171, and Arg173. Although such combination has been observed only in SPAP, these residues are present separately in different members of the AP superfamily. Here, we establish the presence of two distinct classes of APs; ECAP-type and SPAP-type. Bioinformatic analyses show that SPAP-type of APs are widely distributed in the bacterial kingdom. The role of active site residues in the catalytic mechanism has been delineated through a set of crystal structures reported here. These structures, representing different stages of the reaction pathway provide wealth of information for the catalytic mechanism. Despite critical differences in the substrate binding residues, SPAP follows a mechanism similar to that of ECAP-type of APs. Structure-based phylogenetic analysis suggests that SPAP and ECAP may have diverged very early during the evolution from a common ancestor. Moreover, it is proposed that the SPAP-type of APs are fundamental members of the AP superfamily and are more closely related to other members of the superfamily as compared to the ECAP-type of APs.
碱性磷酸酶（APs），以大肠杆菌 AP（ECAP）为代表，利用精氨酸残基稳定活性位点中的磷酸基团；而来自鞘氨醇单胞菌的 AP（SPAP）则表现出独特的底物结合残基组合；Thr89、Asn110、Lys171 和 Arg173。尽管这种组合仅在 SPAP 中观察到，但这些残基在 AP 超家族的不同成员中分别存在。在这里，我们确定了存在两种不同类型的 APs；ECAP 型和 SPAP 型。生物信息学分析表明，SPAP 型 APs 在细菌王国中广泛分布。通过一组这里报道的晶体结构，我们阐明了活性位点残基在催化机制中的作用。这些代表不同反应途径阶段的结构为催化机制提供了丰富的信息。尽管在底物结合残基上存在显著差异，但 SPAP 遵循的机制与 ECAP 型 APs 相似。基于结构的系统发育分析表明，SPAP 和 ECAP 可能在进化过程中很早就从共同祖先分化而来。此外，有人提出 SPAP 型 APs 是 AP 超家族的基本成员，与超家族的其他成员的关系比 ECAP 型 APs 更为密切。
