Quantum-Based Modeling of Dephosphorylation in the Catalytic Site of Serine/Threonine Protein Phosphatase-5 (PPP5C)

Abstract

Serine/threonine protein phosphatase-5 (PP5; PPP5C) is a member of the phosphoprotein phosphatase (PPP) gene family. The PPP catalytic domains feature a bimetal system (M1/M2), an associated bridge hydroxide (W1(OH−)), an M1-bound water/hydroxide (W2), and a highly conserved core sequence. The PPPs are presumed to share a common mechanism: The seryl/threonyl phosphoryl group of the phosphoprotein coordinates the metal ions, W1(OH−) attacks the central phosphorous atom, rupturing the antipodal phosphoester bond and releasing the phosphate-free protein. Also, a histidine/aspartate tandem is responsible for protonating the exiting seryl/threonyl alkoxide. Here, we employed quantum-based computations on a large section of the PP5 catalytic site. A 33-residue, ONIOM(UB3LYP/6-31G(d):UPM7) model was built to perform computations using methylphosphate dianion as a stand-in substrate for phosphoserine/phosphothreonine. We present a concerted transition state (TS) in which W1(OH−) attacks the phosphate center at the same time that the exiting seryl/threonyl alkoxide is protonated directly by the His304/Asp274 tandem, with W2 assigned as a water molecule: W2(H2O). Arg275, proximal to M1, stabilizes the substrate and TS by binding both the ester oxygen (Oγ) and a phosphoryl oxygen (O1) in a bidentate fashion; in the product state, Tyr451 aids in decoupling Arg275 from O1 of the product phosphate ion. The reaction is exothermic (ΔH = −2.0 kcal/mol), occurs in a single step, and has a low activation barrier (ΔH‡ = +10.0 kcal/mol). Our work is an improvement over an earlier computational study that also found bond rupture and alkoxide protonation to be concerted, but concluded that Arg275 is deprotonated during the reactant and TS stages of the pathway. In that earlier study, the critical electron-withdrawal role that Arg275 plays during the hydroxide attack was not correctly accounted for.

Quantum-based modeling implies that bidentate Arg89-substrate binding enhances serine/threonine protein phosphatase-2A(PPP2R5D/PPP2R1A/PPP2CA)-mediated dephosphorylation

“…Using a quantum-based computational model of the PP2A(PPP2R5D)/pSer system prepared from structural data, we have found that when Arg 89 is engaged with Glu 198 in a salt bridge (Pathway II-alt), the energy of activation is higher than when bidentate Arg 89 -pSer binding occurs (Pathway I), Table 1 . Geometric features of the reactant and transition states for Pathway I also support the conclusion that the bidentate motif, established for PPP5 in our earlier study ( Salter et al, 2020 ), is needed for optimal efficiency of PP2A(PPP2R5X) hydrolysis and is likely a universal characteristic of the PPPs.…”

“…Animation of Pathway I’s imaginary mode (441.7 i cm -1 ) confirms that the TS is concerted, just as in PPP5/pMeOH ( Salter et al, 2020 ); that is, W 1 (OH − ) attacks the P center at the same time that the exiting seryl alkoxide is protonated by His 118 . This TS mode differs, however, in that the inversion motion of the P center is less pronounced relative to the motion of H + transfer from His 118 , presumably because the inversion is already well advanced at 10.6° versus 4.1° for PPP5/pMeOH.…”

“…ONIOM QM/QM calculations: The quantum regions from Step 3 were extracted as Models A and B to perform calculations using Gaussian16 ( Frisch et al, 2016 ). ONIOM(UB3LYP/6-31G(d):UPM7) ( Ditchfield, 1971 ; Hehre, 1972 ; Becke, 1993 ; Lee et al, 1998 ; Rassolov et al, 1998 ; Dapprich et al, 1999 ) partial optimizations for stable structures and searches for TS structures were carried out as in Salter et al (2020) . Atoms were selected for the “high-level” and “low-level” regions as indicated in Figure 2 .…”

MiR-520a-5p/PPP5C regulation pattern is identified as the key to gemcitabine resistance in pancreatic cancer

“…If PPP5C is dephosphorylated at the P site, its highly conserved catalytic core and bimetallic system (M1/M2) will change to substrate-binding and hydrolysis sites. This will lead to the overexpression of PPP5C, which is one of the reasons for cell proliferation and the progression of various cancers ( 23 ). As an essential regulator of hormone and stress-related signal transduction ( 24 ), inhibition of its activity or expression will cause cell cycle arrest, hinder mitosis, and eventually lead to apoptosis ( 25 ).…”

Computational Catalysis—Trends and Outlook

“…This Special Issue highlights the diverse impact that computational catalysis investigations can provide, ranging from key reaction systems in the energy sector to complex biological processes [1]. In addition, the rapid evolution of the underlying theoretical methods, such as the unified reaction valley approach highlighted in this Special Issue, provides powerful tools for the broader community.…”