Polar Group Burial Contributes More to Protein Stability than Nonpolar Group Burial

Cn, Pace

doi:10.1021/bi001574j

Cited by 140 publications

(127 citation statements)

References 29 publications

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“…The hRI·RNase 1 complex also buries an additional 375 Å 2 of surface area, which could also enhance its stability. 48,45,49 In line with the more intimate complex formed by hRI and RNase 1, RNase 1 dissociates from hRI 150-fold more slowly than does RNase A from hRI ( Figure 5 ; Table 3). 12 The slower dissociation rate (t ½ = 81 days) for the hRI·RNase 1 complex is in the range of that for the hRI·angiogenin complex (t ½ = 62 days).…”

Section: Recognition Of Rnase 1 By Rimentioning

confidence: 80%

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Inhibition of Human Pancreatic Ribonuclease by the Human Ribonuclease Inhibitor Protein

Johnson

McCoy

Bingman

et al. 2007

Journal of Molecular Biology

135

201

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The ribonuclease inhibitor protein (RI) binds to members of the bovine pancreatic ribonuclease (RNase A) superfamily with an affinity in the femtomolar range. Here, we report on structural and energetic aspects of the interaction between human RI (hRI) and human pancreatic ribonuclease (RNase 1). The structure of the crystalline hRI·RNase 1 complex was determined at a resolution of 1.95 Å, revealing the formation of 19 intermolecular hydrogen bonds involving 13 residues of RNase 1. In contrast, only 9 such hydrogen bonds are apparent in the structure of the complex between porcine RI and RNase A. hRI, which is anionic, also appears to use its horseshoe-shaped structure to engender long-range Coulombic interactions with RNase 1, which is cationic. In accordance with the structural data, the hRI·RNase 1 complex was found to be extremely stable (t ½ = 81 days; K d = 2.9 × 10 -16 M). Site-directed mutagenesis experiments enabled the identification of two cationic residues in RNase 1-Arg39 and Arg91-that are especially important for both the formation and stability of the complex, and are thus termed "electrostatic targeting residues". Disturbing the electrostatic attraction between hRI and RNase 1 yielded a variant of RNase 1 that maintained ribonucleolytic activity and conformational stability but had a 2.8 × 10 3 -fold lower association rate for complex formation and 5.9 × 10 9 -fold lower affinity for hRI. This variant of RNase 1, which exhibits the largest decrease in RI affinity of any engineered ribonuclease, is also toxic to human erythroleukemia cells. Together, these results provide new insight into an unusual and important protein-protein interaction, and could expedite the development of human ribonucleases as chemotherapeutic agents.

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Section: Recognition Of Rnase 1 By Rimentioning

confidence: 80%

“…[43][44][45][46] Hydrogen bonds between functional groups of opposite charge (i.e., salt bridges) are especially strong. 47 Overall, the hRI·RNase 1 complex has both more intermolecular hydrogen bonds than does the pRI·RNase A complex and more between residues of opposite charge (Table 2).…”

Section: Recognition Of Rnase 1 By Rimentioning

confidence: 99%

Inhibition of Human Pancreatic Ribonuclease by the Human Ribonuclease Inhibitor Protein

Johnson

McCoy

Bingman

et al. 2007

Journal of Molecular Biology

135

201

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“…The theoretical studies that reach different conclusions need to be re-examined. We have emphasized previously the importance of van der Waals interactions in the tightly packed interior of a protein to the contribution of polar group burial to stability (4). Recently, several groups have pointed out that the enhanced van der Waals interactions resulting from the tight packing of groups in the interior of folded proteins are also of crucial importance to the hydrophobic effect (24,40,41).…”

Section: Discussionmentioning

confidence: 99%

“…We recently suggested that polar group burial makes a larger contribution to the stability of globular proteins than nonpolar group burial (4). This idea is supported by most experimental studies (5-11), but not by most theoretical studies (12)(13)(14).…”

mentioning

confidence: 97%

The Contribution of Polar Group Burial to Protein Stability Is Strongly Context-dependent

Takano

Scholtz

Sacchettini

et al. 2003

Journal of Biological Chemistry

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We previously suggested that proteins gain more stability from the burial and hydrogen bonding of polar groups than from the burial of nonpolar groups (Pace, C. N. (2001) Biochemistry 40, 310 -313). To study this further, we prepared eight Thr-to-Val mutants of RNase Sa, four in which the Thr side chain is hydrogen-bonded and four in which it is not. We measured the stability of these mutants by analyzing their thermal denaturation curves. The four hydrogen-bonded Thr side chains contribute 1.3 ؎ 0.9 kcal/mol to the stability; those that are not still contribute 0.4 ؎ 0.9 kcal/mol to the stability. For 40 Thr-to-Val mutants of 11 proteins, the average decrease in stability is 1.0 ؎ 1.0 kcal/mol when the Thr side chain is hydrogen-bonded and 0.0 ؎ 0.5 kcal/mol when it is not. This is clear evidence that hydrogen bonds contribute favorably to protein stability. In addition, we prepared four Val-to-Thr mutants of RNase Sa, measured their stability, and determined their crystal structures. In all cases, the mutants are less stable than the wild-type protein, with the decreases in stability ranging from 0.5 to 4.4 kcal/mol. For 41 Val-to-Thr mutants of 11 proteins, the average decrease in stability is 1.8 ؎ 1.3 kcal/mol and is unfavorable for 40 of 41 mutants. This shows that placing an -OH group at a site designed for a -CH 3 group is very unfavorable. So, -OH groups can contribute favorably to protein stability, even if they are not hydrogen-bonded, if the site was selected for an -OH group, but they will make an unfavorable contribution to stability, even if they are hydrogen-bonded, when they are placed at a site selected for a -CH 3 group. The contribution that polar groups make to protein stability depends strongly on their environment.

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“…The formation of secondary structure effectively buries hydrogen bonds, which further decreases the free energy of the emergent protein. In fact, burial of hydrogen bonds or polar groups in the hydrophobic core of a protein contributes more to overall stability than burial of the volume equivalent of non-polar groups [2,3]. As folding progresses, the secondary structure elements are assembled into native-like tertiary structures, where the exclusion of water due to tight packing of the structural elements further decreases the free energy of the nascent protein as it descends through the energy landscape towards its native structure.…”

Section: Tertiary Structure -Hold the Foldmentioning

confidence: 99%

Thiopurine S-methyltransferase - characterization of variants and ligand binding

Blissing¹

2017

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Populärvetenskaplig sammanfattningProteinet TPMT och tiopurinläkemedel I kroppens celler finns proteinet tiopurinmetyltransferas, vilket vanligtvis förkortas TPMT. Trots att det är vanligt förekommande är proteinets naturliga funktion fortfarande okänd. Vi vet dock att TPMT inaktiverar en viss typ av läkemedel som kallas tiopuriner, därav proteinets namn. Tiopuriner är en typ av cytostatika (cellgifter) som använd vid behandling av sjukdomstillstånd relaterade till ett överaktivt immunförsvar och vissa former av leukemi. Tiopurinläkemedel är förstklassiga imitatörer; de efterliknar nämligen byggstenarna till vårt DNA. Cellen kan inte skilja imitatör från original utan bygger in tiopurinerna i nytt DNA. DNA som innehåller tiopuriner är odugligt, vilket gör att cellen till slut dör. Eftersom cellen kopierar sitt DNA före delning så kommer tiopurinläkemedel i större utsträckning påverka celler som delar sig ofta, och man får därmed en riktad effekt mot överaktiva immunceller och cancerceller. TPMT komplicerar läkemedelsbehandlingProteinet TPMT inaktiverar tiopurinläkemedel, och det får tyvärr konsekvenser för patienter som behandlas med dessa substanser. Inaktiveringen sker genom att proteinet modifierar tiopurinerna genom så kallad metylering. De modifierade läkemedelsmolekylerna liknar inte längre de naturliga DNA-baserna, och läkemedlets celldödande effekt minskar därmed. För att kompensera för TPMTs härjningar måste läkemedelsdosen höjas så att tillräcklig effekt uppnås. Dock finns det flertalet naturligt förekommande varianter av proteinet, vilket ytterligare komplicerar behandlingen. Att det finns olika varianter av ett protein är en naturlig i del av den genetiska variationen i en population, men alla TPMTvarianterna inte är lika benägna att inaktivera tiopurinläkemedel. Vissa varianter är ambitiösa och inaktiverar en större mängd tiopurinerde är alltså högaktiva. Andra är betydligt sämre på att inaktivera läkemedelsmolekyler -de är med andra ord lågaktiva. En konsekvens av varianternas olika egenskaper är att det förekommer individuella skillnader i hur man reagerar på tiopurinläkemedel, beroende på vilken typ av TPMT-protein man har anlag för. Det gör doseringen av tiopuriner komplicerad. En patient som har gener för en lågaktiv TPMT-variant riskerar att överdoseras, vilket kan leda till att immunförsvaret slås ut helt. En patient som har högaktivt TPMT-protein bildar istället stora mängder inaktiverade läkemedelsmolekyler, vilka kan skada levern vid höga halter. För att göra behandling med tiopurinläkemedel säkrare görs rutinmässigt en kontroll av vilken typ av TPMT-protein patienten har, och därefter skräddarsys doseringen för varje patient. Man övervakar också patienten med regelbundna provtagningar för att kontrollera att mängden läkemedel är på rätt nivå. Biokemin som hjälpmedelI vår forskning studerar vi utvalda TPMT-varianter för att förklara vad som orsakar deras olika biofysikaliska egenskaper. Vi har bland annat kunnat visa att den molekylära strukturen hos en variant, TPMT*6 (Y180F), är lite min...

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Polar Group Burial Contributes More to Protein Stability than Nonpolar Group Burial

Cited by 140 publications

References 29 publications

Inhibition of Human Pancreatic Ribonuclease by the Human Ribonuclease Inhibitor Protein

Inhibition of Human Pancreatic Ribonuclease by the Human Ribonuclease Inhibitor Protein

The Contribution of Polar Group Burial to Protein Stability Is Strongly Context-dependent

Thiopurine S-methyltransferase - characterization of variants and ligand binding

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