Phosphoribosyl Pyrophosphate Synthetase 1

Grateful Beckers and Maya Vaccaro '24

Contents:

I. Introduction

Human Phosphoribosyl Pyrophosphate Synthetase 1 (hPRS1) is an enzyme that plays an intermediate role in building new nucleotides, as well as NAD, NADP, histidine, and tryptophan. Specifically, it catalyzes a reaction that transfers two phosphates from an ATP to Ribose 5-Phosphate (R5P) yielding Phosphoribosyl Pyrophosphate (PRPP). As a result of their importance in the production of essential biomolecules, phosphoribosyl Pyrophosphate synthetases are expressed constitutively among most organisms. There are three major classes of this protein. Most PRSs, including hPRS, fall into the category of Class I. These PRSs interact with Mg and Phosphate ions in order to catalyze PRPP production, and are allosterically inhibited by ADP. Class II PRSs are found only in plants. They do not require phosphate ions in order to catalyze the production of PRPP, can use a larger array of nTPs to catalyze PRPP production, and lack an allosteric binding site for ADP. Class III PRSs are found in the archaea Methanocaldococcus jannaschii. While this class of PRS does rely on phosphate ions for activation, it also lacks an ADP allosteric site.

II. General Structure

hPRS1 forms a in its functional state. There are three asymmetric units in this hexamer, each made up of two PRS1 proteins bound together to form a with the first subunit containing 305 amino acids, Asn3 to Phe313 except for Lys197 to Val202, while the second consists of 308 amino acids, Asn3 to Pro317 except for Arg196 to Val202. Each asymmetric unit is associated with six phosphate ions and . The binding sites for ATP and R5P are located at an active site where these two domains of the homodimer meet in each subunit. The previously established allosteric site, where phosphate and ADP bind, is located at the interface of three subunits of the hexamer, and a recently discovered allosteric site is located between each active site and previously established allosteric site.

III. Active Site

The active site of hPRS1 is composed of the binding sites where ATP and R5P bind in order to facilitate the production of PRPP. There is an ATP binding site on each interface of the two asymmetric subunits of the hexamer. There are three main parts to the binding site. The is located on Phe92 to Ser108 of one subunit in the dimer, the loop is located on Asp171 to Gly174 of the same subunit, and the located on Val30 to Ile44 of the other subunit in the dimer. This active site forms a complex with ATP, SO42- and Cd2+. The AMP portion of ATP interacts with Arg96, Gln97, Asp101 and His130 of one subunit and Phe35, Asn37 and Glu39 of another subunit. R5P binds to residues Asp220 through Thr228 of each subunit. R5P forms a complex with sulfate and cadmium, with cadmium occupying the same place as the 5-phosphate of R5P. This sulfate has hydrogen bonding interactions with the main chains of Asp224, Thr225, Cys226, Gly227 and the side chains of Thr225 and Thr228.

IV.Allosteric Binding

There are two known allosteric binding sites on hPRS1. The more well-studied allosteric site is located across three subunits of the hexameric structure. Each is composed of residues Ser135, Asp143, Asn144 and Ser308 through Phe313 of the first subunit, Lys100 through Arg104 of the flexible loop, and Ser47, Arg49, Ala80 and Ser81 of the third subunit. Phosphate or sulfate binds as an allosteric activator. Binding at this allosteric site leads to stabilizing the of the protein, fixing the protein in the open, active conformation. This open conformation allows ATP and R5P to bind at the active site, and synthesis of PRPP to commence.The bound phosphate (or sulfate) also has hydrogen bonding interactions with the side chains of Ser47, Arg49, Arg104, Ser308 and Ser310, the main chains of Val109 and Ser310 , and two water molecules. This open conformation allows ATP and R5P to bind at the active site, and synthesis of PRPP to commence. ADP is an allosteric inhibitor that competes with phosphate for the allosteric site.

A was found between the active site and the first allosteric site. A single phosphate (or, in the case of the study which uncovered this allosteric site, a sulfate) binds, stabilizing the flexible loop. In this binding site, the phosphate/sulfate forms hydrogen bonds with the side chains of Ser132, Gln135, Asn144 and Tyr146of one subunit in the dimer, and Lys100, Asp101 and Lys102 of the flexible loop of the other subunit in the dimer. Binding at this site also repositions the flexible loop so that ADP cannot bind to the first allosteric site, preventing the allosteric inhibition of the reaction. Conversely, the binding of ADP to the first allosteric site prevents the binding of a phosphate/sulfate to the second allosteric site.

VI. References

Chen P., Liu Z., Wang X., Peng J., Sun, Q., Li J., Wang M., Niu L., Zhang Z., Cai G., Teng T., Li X.. (2015). Crystal and EM Structures of Human Phosphoribosyl Pyrophosphate Synthase I (PRS1) Provide Novel Insights into the Disease-Associated Mutations. PLOS ONE, Public Library of Science. https://doi.org/10.1371/journal.pone.0120304

Li S., Lu Y., Peng B., Ding J.. (2007). Crystal Structure of Human Phosphoribosylpyrophosphate Synthetase 1 Reveals a Novel Allosteric Site. The Biochemical Journal, Portland Press Ltd., 1 401(Pt 1): 39 through 47. 10.1042/BJ20061066

Hove-Jensen, B., Anderson K. R., Kilstrup M., Martinussen J., Switzer R. L., Willemoem. (2016). Phosphoribosyl Diphosphate (PRPP): Biosynthesis, Enzymology, Utilization, and Metabolic Significance. Microbiology and Molecular Biology Reviews : MMBR, U.S. National Library of Medicine. 81(1). 10.1128/MMBR.00040-16 28:4568 through 4574.

Schneiter, Roger, et al. The Importance of the Five Phosphoribosyl-Pyrophosphate Synthetase (PRS) Gene Products of Saccharomyces Cerevisiae in the Maintenance of Cell Integrity and the Subcellular Localization of PRS1P. Microbiology, Microbiology Society, 1 Dec. 2000, https://www.microbiologyresearch.org/content/journal/micro/10.1099/00221287-146-12-3269..

Tong X., Zhao F., Thompson B .. (2009) The Molecular Determinants of De Novo Nucleotide Biosynthesis in Cancer Cells. Current Opinion in Genetics & Development, Elsevier Current Trends. 19(1). 32-37. https://doi.org/10.1016/j.gde.2009.01.002

Wood, A. W., and Seegmiller E. J.. ( 2021) Properties of 5-Phosphoribosyl-1-Pyrophosphate Amidotransferase from Human Lymphoblasts. Journal of Biological Chemistry. 248(1). 138 through 143. https://doi.org/10.1016/S0021-9258(19)44455-4.

Back to Top