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Articles 1 - 30 of 36
Full-Text Articles in Biochemistry, Biophysics, and Structural Biology
A Sensitive Assay Using A Native Protein Substrate For Screening Hiv-1 Maturation Inhibitors Targeting The Protease Cleavage Site Between The Matrix And Capsid, Sook-Kyung Lee, Nancy Cheng, Emily Hull-Ryde, Marc Potempa, Celia Schiffer, William Janzen, Ronald Swanstrom
A Sensitive Assay Using A Native Protein Substrate For Screening Hiv-1 Maturation Inhibitors Targeting The Protease Cleavage Site Between The Matrix And Capsid, Sook-Kyung Lee, Nancy Cheng, Emily Hull-Ryde, Marc Potempa, Celia Schiffer, William Janzen, Ronald Swanstrom
Celia A. Schiffer
The matrix/capsid processing site in the HIV-1 Gag precursor is likely the most sensitive target to inhibit HIV-1 replication. We have previously shown that modest incomplete processing at the site leads to a complete loss of virion infectivity. In the study presented here, a sensitive assay based on fluorescence polarization that can monitor cleavage at the MA/CA site in the context of the folded protein substrate is described. The substrate, an MA/CA fusion protein, was labeled with the fluorescein-based FlAsH (fluorescein arsenical hairpin) reagent that binds to a tetracysteine motif (CCGPCC) that was introduced within the N-terminal domain of CA. …
Interview With Celia Schiffer, Celia Schiffer
Interview With Celia Schiffer, Celia Schiffer
Celia A. Schiffer
Celia Schiffer, a Professor in Biochemistry and Molecular Pharmacology; a former Director of UMass Center for AIDS Research; and a Founder and Co-Director for the Institute for Drug Resistance (University of Massachusetts Medical School, MA, USA). Schiffer has an undergraduate degree in physics from the University of Chicago, with a PhD in biophysics from University of California, San Francisco (CA, USA). She was a postdoctoral associate first at the ETH in Zurich and then at Genentech in San Francisco. Schiffer has published more than 100 peer reviewed journal articles. Her laboratory primarily uses structural biology, biophysical and chemistry techniques to …
Substrate Envelope-Designed Potent Hiv-1 Protease Inhibitors To Avoid Drug Resistance, Madhavi Nalam, Akbar Ali, G. S. Kiran Kumar Reddy, Hong Cao, Saima Anjum, Michael Altman, Nese Yilmaz, Bruce Tidor, Tariq Rana, Celia Schiffer
Substrate Envelope-Designed Potent Hiv-1 Protease Inhibitors To Avoid Drug Resistance, Madhavi Nalam, Akbar Ali, G. S. Kiran Kumar Reddy, Hong Cao, Saima Anjum, Michael Altman, Nese Yilmaz, Bruce Tidor, Tariq Rana, Celia Schiffer
Celia A. Schiffer
The rapid evolution of HIV under selective drug pressure has led to multidrug resistant (MDR) strains that evade standard therapies. We designed highly potent HIV-1 protease inhibitors (PIs) using the substrate envelope model, which confines inhibitors within the consensus volume of natural substrates, providing inhibitors less susceptible to resistance because a mutation affecting such inhibitors will simultaneously affect viral substrate processing. The designed PIs share a common chemical scaffold but utilize various moieties that optimally fill the substrate envelope, as confirmed by crystal structures. The designed PIs retain robust binding to MDR protease variants and display exceptional antiviral potencies against …
Cooperative Effects Of Drug-Resistance Mutations In The Flap Region Of Hiv-1 Protease, Jennifer Foulkes-Murzycki, Christina Rosi, Nese Yilmaz, Robert Shafer, Celia Schiffer
Cooperative Effects Of Drug-Resistance Mutations In The Flap Region Of Hiv-1 Protease, Jennifer Foulkes-Murzycki, Christina Rosi, Nese Yilmaz, Robert Shafer, Celia Schiffer
Celia A. Schiffer
Understanding the interdependence of multiple mutations in conferring drug resistance is crucial to the development of novel and robust inhibitors. As HIV-1 protease continues to adapt and evade inhibitors while still maintaining the ability to specifically recognize and efficiently cleave its substrates, the problem of drug resistance has become more complicated. Under the selective pressure of therapy, correlated mutations accumulate throughout the enzyme to compromise inhibitor binding, but characterizing their energetic interdependency is not straightforward. A particular drug resistant variant (L10I/G48V/I54V/V82A) displays extreme entropy-enthalpy compensation relative to wild-type enzyme but a similar variant (L10I/G48V/I54A/V82A) does not. Individual mutations of sites …
Structural And Thermodynamic Basis Of Amprenavir/Darunavir And Atazanavir Resistance In Hiv-1 Protease With Mutations At Residue 50, Seema Mittal, Rajintha Bandaranayake, Nancy King, Moses Prabu-Jeyabalan, Madhavi Nalam, Ellen Nalivaika, Nese Yilmaz, Celia Schiffer
Structural And Thermodynamic Basis Of Amprenavir/Darunavir And Atazanavir Resistance In Hiv-1 Protease With Mutations At Residue 50, Seema Mittal, Rajintha Bandaranayake, Nancy King, Moses Prabu-Jeyabalan, Madhavi Nalam, Ellen Nalivaika, Nese Yilmaz, Celia Schiffer
Celia A. Schiffer
Drug resistance occurs through a series of subtle changes that maintain substrate recognition but no longer permit inhibitor binding. In HIV-1 protease, mutations at I50 are associated with such subtle changes that confer differential resistance to specific inhibitors. Residue I50 is located at the protease flap tips, closing the active site upon ligand binding. Under selective drug pressure, I50V/L substitutions emerge in patients, compromising drug susceptibility and leading to treatment failure. The I50V substitution is often associated with amprenavir (APV) and darunavir (DRV) resistance, while the I50L substitution is observed in patients failing atazanavir (ATV) therapy. To explain how APV, …
Extreme Entropy-Enthalpy Compensation In A Drug-Resistant Variant Of Hiv-1 Protease, Nancy King, Moses Prabu-Jeyabalan, Rajintha Bandaranayake, Madhavi Nalam, Ellen Nalivaika, Aysegul Ozen, Turkan Haliloglu, Nese Yilmaz, Celia Schiffer
Extreme Entropy-Enthalpy Compensation In A Drug-Resistant Variant Of Hiv-1 Protease, Nancy King, Moses Prabu-Jeyabalan, Rajintha Bandaranayake, Madhavi Nalam, Ellen Nalivaika, Aysegul Ozen, Turkan Haliloglu, Nese Yilmaz, Celia Schiffer
Celia A. Schiffer
The development of HIV-1 protease inhibitors has been the historic paradigm of rational structure-based drug design, where structural and thermodynamic analyses have assisted in the discovery of novel inhibitors. While the total enthalpy and entropy change upon binding determine the affinity, often the thermodynamics are considered in terms of inhibitor properties only. In the current study, profound changes are observed in the binding thermodynamics of a drug-resistant variant compared to wild-type HIV-1 protease, irrespective of the inhibitor bound. This variant (Flap+) has a combination of flap and active site mutations and exhibits extremely large entropy-enthalpy compensation compared to wild-type protease, …
Decomposing The Energetic Impact Of Drug-Resistant Mutations: The Example Of Hiv-1 Protease-Drv Binding, Yufeng Cai, Celia Schiffer
Decomposing The Energetic Impact Of Drug-Resistant Mutations: The Example Of Hiv-1 Protease-Drv Binding, Yufeng Cai, Celia Schiffer
Celia A. Schiffer
HIV-1 protease is a major drug target for AIDS therapy. With the appearance of drug-resistant HIV-1 protease variants, understanding the mechanism of drug resistance becomes critical for rational drug design. Computational methods can provide more details about inhibitor-protease binding than crystallography and isothermal titration calorimetry. The latest FDA-approved HIV-1 protease inhibitor is Darunavir (DRV). Herein, each DRV atom is evaluated by free energy component analysis for its contribution to the binding affinity with wild-type protease and ACT, a drug-resistant variant. This information can contribute to the rational design of new HIV-1 protease inhibitors.
Context Surrounding Processing Sites Is Crucial In Determining Cleavage Rate Of A Subset Of Processing Sites In Hiv-1 Gag And Gag-Pro-Pol Polyprotein Precursors By Viral Protease, Sook-Kyung Lee, Marc Potempa, Madhavi Kolli, Aysegul Ozen, Celia Schiffer, Ronald Swanstrom
Context Surrounding Processing Sites Is Crucial In Determining Cleavage Rate Of A Subset Of Processing Sites In Hiv-1 Gag And Gag-Pro-Pol Polyprotein Precursors By Viral Protease, Sook-Kyung Lee, Marc Potempa, Madhavi Kolli, Aysegul Ozen, Celia Schiffer, Ronald Swanstrom
Celia A. Schiffer
Processing of the human immunodeficiency virus type 1 (HIV-1) Gag and Gag-Pro-Pol polyproteins by the HIV-1 protease (PR) is essential for the production of infectious particles. However, the determinants governing the rates of processing of these substrates are not clearly understood. We studied the effect of substrate context on processing by utilizing a novel protease assay in which a substrate containing HIV-1 matrix (MA) and the N-terminal domain of capsid (CA) is labeled with a FlAsH (fluorescein arsenical hairpin) reagent. When the seven cleavage sites within the Gag and Gag-Pro-Pol polyproteins were placed at the MA/CA site, the rates of …
Structural, Kinetic, And Thermodynamic Studies Of Specificity Designed Hiv-1 Protease, Oscar Alvizo, Seema Mittal, Stephen Mayo, Celia Schiffer
Structural, Kinetic, And Thermodynamic Studies Of Specificity Designed Hiv-1 Protease, Oscar Alvizo, Seema Mittal, Stephen Mayo, Celia Schiffer
Celia A. Schiffer
HIV-1 protease recognizes and cleaves more than 12 different substrates leading to viral maturation. While these substrates share no conserved motif, they are specifically selected for and cleaved by protease during viral life cycle. Drug resistant mutations evolve within the protease that compromise inhibitor binding but allow the continued recognition of all these substrates. While the substrate envelope defines a general shape for substrate recognition, successfully predicting the determinants of substrate binding specificity would provide additional insights into the mechanism of altered molecular recognition in resistant proteases. We designed a variant of HIV protease with altered specificity using positive computational …
Design, Synthesis, And Biological And Structural Evaluations Of Novel Hiv-1 Protease Inhibitors To Combat Drug Resistance, Maloy Parai, David Huggins, Hong Cao, Madhavi Nalam, Akbar Ali, Celia Schiffer, Bruce Tidor, Tariq Rana
Design, Synthesis, And Biological And Structural Evaluations Of Novel Hiv-1 Protease Inhibitors To Combat Drug Resistance, Maloy Parai, David Huggins, Hong Cao, Madhavi Nalam, Akbar Ali, Celia Schiffer, Bruce Tidor, Tariq Rana
Celia A. Schiffer
A series of new HIV-1 protease inhibitors (PIs) were designed using a general strategy that combines computational structure-based design with substrate-envelope constraints. The PIs incorporate various alcohol-derived P2 carbamates with acyclic and cyclic heteroatomic functionalities into the (R)-hydroxyethylamine isostere. Most of the new PIs show potent binding affinities against wild-type HIV-1 protease and three multidrug resistant (MDR) variants. In particular, inhibitors containing the 2,2-dichloroacetamide, pyrrolidinone, imidazolidinone, and oxazolidinone moieties at P2 are the most potent with K(i) values in the picomolar range. Several new PIs exhibit nanomolar antiviral potencies against patient-derived wild-type viruses from HIV-1 clades A, B, and C …
Hydrophobic Core Flexibility Modulates Enzyme Activity In Hiv-1 Protease, Seema Mittal, Yufeng Cai, Madhavi Nalam, Daniel Bolon, Celia Schiffer
Hydrophobic Core Flexibility Modulates Enzyme Activity In Hiv-1 Protease, Seema Mittal, Yufeng Cai, Madhavi Nalam, Daniel Bolon, Celia Schiffer
Celia A. Schiffer
Human immunodeficiency virus Type-1 (HIV-1) protease is crucial for viral maturation and infectivity. Studies of protease dynamics suggest that the rearrangement of the hydrophobic core is essential for enzyme activity. Many mutations in the hydrophobic core are also associated with drug resistance and may modulate the core flexibility. To test the role of flexibility in protease activity, pairs of cysteines were introduced at the interfaces of flexible regions remote from the active site. Disulfide bond formation was confirmed by crystal structures and by alkylation of free cysteines and mass spectrometry. Oxidized and reduced crystal structures of these variants show the …
Evaluation Of The Substrate Envelope Hypothesis For Inhibitors Of Hiv-1 Protease, Sripriya Chellappan, Visvaldas Kairys, Miguel Fernandes, Celia Schiffer, Michael Gilson
Evaluation Of The Substrate Envelope Hypothesis For Inhibitors Of Hiv-1 Protease, Sripriya Chellappan, Visvaldas Kairys, Miguel Fernandes, Celia Schiffer, Michael Gilson
Celia A. Schiffer
Crystallographic data show that various substrates of HIV protease occupy a remarkably uniform region within the binding site; this region has been termed the substrate envelope. It has been suggested that an inhibitor that fits within the substrate envelope should tend to evade viral resistance because a protease mutation that reduces the affinity of the inhibitor will also tend to reduce the affinity of substrate, and will hence decrease the activity of the enzyme. Accordingly, inhibitors that fit the substrate envelope better should be less susceptible to clinically observed resistant mutations, since these must also allow substrates to bind. The …
Discovery Of Hiv-1 Protease Inhibitors With Picomolar Affinities Incorporating N-Aryl-Oxazolidinone-5-Carboxamides As Novel P2 Ligands, Akbar Ali, G. S. Kiran Kumar Reddy, Hong Cao, Saima Anjum, Madhavi Nalam, Celia Schiffer, Tariq Rana
Discovery Of Hiv-1 Protease Inhibitors With Picomolar Affinities Incorporating N-Aryl-Oxazolidinone-5-Carboxamides As Novel P2 Ligands, Akbar Ali, G. S. Kiran Kumar Reddy, Hong Cao, Saima Anjum, Madhavi Nalam, Celia Schiffer, Tariq Rana
Celia A. Schiffer
Here, we describe the design, synthesis, and biological evaluation of novel HIV-1 protease inhibitors incorporating N-phenyloxazolidinone-5-carboxamides into the (hydroxyethylamino)sulfonamide scaffold as P2 ligands. Series of inhibitors with variations at the P2 phenyloxazolidinone and the P2' phenylsulfonamide moieties were synthesized. Compounds with the (S)-enantiomer of substituted phenyloxazolidinones at P2 show highly potent inhibitory activities against HIV-1 protease. The inhibitors possessing 3-acetyl, 4-acetyl, and 3-trifluoromethyl groups at the phenyl ring of the oxazolidinone fragment are the most potent in each series, with K(i) values in the low picomolar (pM) range. The electron-donating groups 4-methoxy and 1,3-dioxolane are preferred at P2' phenyl ring, …
Association Of A Novel Human Immunodeficiency Virus Type 1 Protease Substrate Cleft Mutation, L23i, With Protease Inhibitor Therapy And In Vitro Drug Resistance, Elizabeth Johnston, Mark Winters, Soo-Yon Rhee, Thomas Merigan, Celia Schiffer, Robert Shafer
Association Of A Novel Human Immunodeficiency Virus Type 1 Protease Substrate Cleft Mutation, L23i, With Protease Inhibitor Therapy And In Vitro Drug Resistance, Elizabeth Johnston, Mark Winters, Soo-Yon Rhee, Thomas Merigan, Celia Schiffer, Robert Shafer
Celia A. Schiffer
We observed a previously uncharacterized mutation in the protease substrate cleft, L23I, in 31 of 4,303 persons undergoing human immunodeficiency virus type 1 genotypic resistance testing. In combination with V82I, L23I was associated with a sevenfold reduction in nelfinavir susceptibility and a decrease in replication capacity. In combination with other drug resistance mutations, L23I was associated with multidrug resistance and a compensatory increase in replication capacity.
Co-Evolution Of Nelfinavir-Resistant Hiv-1 Protease And The P1-P6 Substrate, Madhavi Kolli, Stephane Lastere, Celia Schiffer
Co-Evolution Of Nelfinavir-Resistant Hiv-1 Protease And The P1-P6 Substrate, Madhavi Kolli, Stephane Lastere, Celia Schiffer
Celia A. Schiffer
The selective pressure of the competitive protease inhibitors causes both HIV-1 protease and occasionally its substrates to evolve drug resistance. We hypothesize that this occurs particularly in substrates that protrude beyond the substrate envelope and contact residues that mutate in response to a particular protease inhibitor. To validate this hypothesis, we analyzed substrate and protease sequences for covariation. Using the chi2 test, we show a positive correlation between the nelfinavir-resistant D30N/N88D protease mutations and mutations at the p1-p6 cleavage site as compared to the other cleavage sites. Both nelfinavir and the substrate p1-p6 protrude beyond the substrate envelope and contact …
Substrate Shape Determines Specificity Of Recognition For Hiv-1 Protease: Analysis Of Crystal Structures Of Six Substrate Complexes, Moses Prabu-Jeyabalan, Ellen Nalivaika, Celia Schiffer
Substrate Shape Determines Specificity Of Recognition For Hiv-1 Protease: Analysis Of Crystal Structures Of Six Substrate Complexes, Moses Prabu-Jeyabalan, Ellen Nalivaika, Celia Schiffer
Celia A. Schiffer
The homodimeric HIV-1 protease is the target of some of the most effective antiviral AIDS therapy, as it facilitates viral maturation by cleaving ten asymmetric and nonhomologous sequences in the Gag and Pol polyproteins. Since the specificity of this enzyme is not easily determined from the sequences of these cleavage sites alone, we solved the crystal structures of complexes of an inactive variant (D25N) of HIV-1 protease with six peptides that correspond to the natural substrate cleavage sites. When the protease binds to its substrate and buries nearly 1000 A2 of surface area, the symmetry of the protease is broken, …
Rationale For More Diverse Inhibitors In Competition With Substrates In Hiv-1 Protease, Nevra Ozer, Celia Schiffer, Turkan Haliloglu
Rationale For More Diverse Inhibitors In Competition With Substrates In Hiv-1 Protease, Nevra Ozer, Celia Schiffer, Turkan Haliloglu
Celia A. Schiffer
The structural fluctuations of HIV-1 protease in interaction with its substrates versus inhibitors were analyzed using the anisotropic network model. The directions of fluctuations in the most cooperative functional modes differ mainly around the dynamically key regions, i.e., the hinge axes, which appear to be more flexible in substrate complexes. The flexibility of HIV-1 protease is likely optimized for the substrates' turnover, resulting in substrate complexes being dynamic. In contrast, in an inhibitor complex, the inhibitor should bind and lock down to inactivate the active site. Protease and ligands are not independent. Substrates are also more flexible than inhibitors and …
Substrate Specificity In Hiv-1 Protease By A Biased Sequence Search Method, Nevra Ozer, Turkan Haliloglu, Celia Schiffer
Substrate Specificity In Hiv-1 Protease By A Biased Sequence Search Method, Nevra Ozer, Turkan Haliloglu, Celia Schiffer
Celia A. Schiffer
Drug resistance in HIV-1 protease can also occasionally confer a change in the substrate specificity. Through the use of computational techniques, a relationship can be determined between the substrate sequence and three-dimensional structure of HIV-1 protease, and be utilized to predict substrate specificity. In this study, we introduce a biased sequence search threading (BSST) methodology to analyze the preferences of substrate positions and correlations between them that might also identify which positions within known substrates can likely tolerate sequence variability and which cannot. The potential sequence space was efficiently explored using a low-resolution knowledge-based scoring function. The low-energy substrate sequences …
Additivity In The Analysis And Design Of Hiv Protease Inhibitors, Robert Jorissen, G. S. Kiran Kumar Reddy, Akbar Ali, Michael Altman, Sripriya Chellappan, Saima Anjum, Bruce Tidor, Celia Schiffer, Tariq Rana, Michael Gilson
Additivity In The Analysis And Design Of Hiv Protease Inhibitors, Robert Jorissen, G. S. Kiran Kumar Reddy, Akbar Ali, Michael Altman, Sripriya Chellappan, Saima Anjum, Bruce Tidor, Celia Schiffer, Tariq Rana, Michael Gilson
Celia A. Schiffer
We explore the applicability of an additive treatment of substituent effects to the analysis and design of HIV protease inhibitors. Affinity data for a set of inhibitors with a common chemical framework were analyzed to provide estimates of the free energy contribution of each chemical substituent. These estimates were then used to design new inhibitors whose high affinities were confirmed by synthesis and experimental testing. Derivations of additive models by least-squares and ridge-regression methods were found to yield statistically similar results. The additivity approach was also compared with standard molecular descriptor-based QSAR; the latter was not found to provide superior …
How Does A Symmetric Dimer Recognize An Asymmetric Substrate? A Substrate Complex Of Hiv-1 Protease, Moses Prabu-Jeyabalan, Ellen Nalivaika, Celia Schiffer
How Does A Symmetric Dimer Recognize An Asymmetric Substrate? A Substrate Complex Of Hiv-1 Protease, Moses Prabu-Jeyabalan, Ellen Nalivaika, Celia Schiffer
Celia A. Schiffer
The crystal structure of an actual HIV-1 protease-substrate complex is presented at 2.0 A resolution (R-value of 19.7 % (R(free) 23.3 %)) between an inactive variant (D25N) of HIV-1 protease and a long substrate peptide, Lys-Ala-Arg-Val-Leu-Ala-Glu-Ala-Met-Ser, which covers a full binding epitope of capsid(CA)-p2, cleavage site. The substrate peptide is asymmetric in both size and charge distribution. To accommodate this asymmetry the two protease monomers adopt different conformations burying a total of 1038 A(2) of surface area at the protease-substrate interface. The specificity for the CA-p2 substrate peptide is mainly hydrophobic, as most of the hydrogen bonds are made with …
Toward The Design Of Mutation-Resistant Enzyme Inhibitors: Further Evaluation Of The Substrate Envelope Hypothesis, Visvaldas Kairys, Michael Gilson, Viney Lather, Celia Schiffer, Miguel Fernandes
Toward The Design Of Mutation-Resistant Enzyme Inhibitors: Further Evaluation Of The Substrate Envelope Hypothesis, Visvaldas Kairys, Michael Gilson, Viney Lather, Celia Schiffer, Miguel Fernandes
Celia A. Schiffer
Previous studies have shown the usefulness of the substrate envelope concept in the analysis and prediction of drug resistance profiles for human immunodeficiency virus protease mutants. This study tests its applicability to several other therapeutic targets: Abl kinase, chitinase, thymidylate synthase, dihydrofolate reductase, and neuraminidase. For the targets where many (> or =6) mutation data are available to compute the average mutation sensitivity of inhibitors, the total volume of an inhibitor molecule that projects outside the substrate envelope V(out), is found to correlate with average mutation sensitivity. Analysis of a locally computed volume suggests that the same correlation would hold …
Dynamics Of Preferential Substrate Recognition In Hiv-1 Protease: Redefining The Substrate Envelope, Aysegul Ozen, Turkan Haliloglu, Celia Schiffer
Dynamics Of Preferential Substrate Recognition In Hiv-1 Protease: Redefining The Substrate Envelope, Aysegul Ozen, Turkan Haliloglu, Celia Schiffer
Celia A. Schiffer
Human immunodeficiency virus type 1 (HIV-1) protease (PR) permits viral maturation by processing the gag and gag-pro-pol polyproteins. HIV-1 PR inhibitors (PIs) are used in combination antiviral therapy but the emergence of drug resistance has limited their efficacy. The rapid evolution of HIV-1 necessitates consideration of drug resistance in novel drug design. Drug-resistant HIV-1 PR variants no longer inhibited efficiently, continue to hydrolyze the natural viral substrates. Though highly diverse in sequence, the HIV-1 PR substrates bind in a conserved three-dimensional shape we termed the substrate envelope. Earlier, we showed that resistance mutations arise where PIs protrude beyond the substrate …
N88d Facilitates The Co-Occurrence Of D30n And L90m And The Development Of Multidrug Resistance In Hiv Type 1 Protease Following Nelfinavir Treatment Failure, Yumi Mitsuya, Mark Winters, W. Jeffrey Fessel, Soo-Yon Rhee, Leo Hurley, Michael Horberg, Celia Schiffer, Andrew Zolopa, Robert Shafer
N88d Facilitates The Co-Occurrence Of D30n And L90m And The Development Of Multidrug Resistance In Hiv Type 1 Protease Following Nelfinavir Treatment Failure, Yumi Mitsuya, Mark Winters, W. Jeffrey Fessel, Soo-Yon Rhee, Leo Hurley, Michael Horberg, Celia Schiffer, Andrew Zolopa, Robert Shafer
Celia A. Schiffer
Nelfinavir was once one of the most commonly used protease inhibitors (PIs). To investigate the genetic mechanisms of multidrug resistance in protease isolates with the primary nelfinavir resistance mutation D30N, we analyzed patterns of protease mutations in 582 viruses with D30N from 460 persons undergoing HIV-1 genotypic resistance testing at Stanford University Hospital from 1997 to 2005. Three patterns of mutational associations were identified. First, D30N was positively associated with N88D but negatively associated with N88S. Second, D30N and L90M were negatively associated except in the presence of N88D, which facilitated the co-occurrence of D30N and L90M. Third, D30N+N88D+L90M formed …
Cooperative Fluctuations Of Unliganded And Substrate-Bound Hiv-1 Protease: A Structure-Based Analysis On A Variety Of Conformations From Crystallography And Molecular Dynamics Simulations, Nese Kurt, Walter Scott, Celia Schiffer, Turkan Haliloglu
Cooperative Fluctuations Of Unliganded And Substrate-Bound Hiv-1 Protease: A Structure-Based Analysis On A Variety Of Conformations From Crystallography And Molecular Dynamics Simulations, Nese Kurt, Walter Scott, Celia Schiffer, Turkan Haliloglu
Celia A. Schiffer
The dynamics of HIV-1 protease, both in unliganded and substrate-bound forms have been analyzed by using an analytical method, Gaussian network model (GNM). The method is applied to different conformations accessible to the protein backbone in the native state, observed in crystal structures and snapshots from fully atomistic molecular dynamics (MD) simulation trajectories. The modes of motion obtained from GNM on different conformations of HIV-1 protease are conserved throughout the MD simulations. The flaps and 40's loop of the unliganded HIV-1 protease structure are identified as the most mobile regions. However, in the liganded structure these flaps lose mobility, and …
Design Of Mutation-Resistant Hiv Protease Inhibitors With The Substrate Envelope Hypothesis, Sripriya Chellappan, G. S. Kiran Kumar Reddy, Akbar Ali, Madhavi Nalam, Saima Anjum, Hong Cao, Visvaldas Kairys, Miguel Fernandes, Michael Altman, Bruce Tidor, Tariq Rana, Celia Schiffer, Michael Gilson
Design Of Mutation-Resistant Hiv Protease Inhibitors With The Substrate Envelope Hypothesis, Sripriya Chellappan, G. S. Kiran Kumar Reddy, Akbar Ali, Madhavi Nalam, Saima Anjum, Hong Cao, Visvaldas Kairys, Miguel Fernandes, Michael Altman, Bruce Tidor, Tariq Rana, Celia Schiffer, Michael Gilson
Celia A. Schiffer
There is a clinical need for HIV protease inhibitors that can evade resistance mutations. One possible approach to designing such inhibitors relies upon the crystallographic observation that the substrates of HIV protease occupy a rather constant region within the binding site. In particular, it has been hypothesized that inhibitors which lie within this region will tend to resist clinically relevant mutations. The present study offers the first prospective evaluation of this hypothesis, via computational design of inhibitors predicted to conform to the substrate envelope, followed by synthesis and evaluation against wild-type and mutant proteases, as well as structural studies of …
Computational Design And Experimental Study Of Tighter Binding Peptides To An Inactivated Mutant Of Hiv-1 Protease, Michael Altman, Ellen Nalivaika, Moses Prabu-Jeyabalan, Celia Schiffer, Bruce Tidor
Computational Design And Experimental Study Of Tighter Binding Peptides To An Inactivated Mutant Of Hiv-1 Protease, Michael Altman, Ellen Nalivaika, Moses Prabu-Jeyabalan, Celia Schiffer, Bruce Tidor
Celia A. Schiffer
Drug resistance in HIV-1 protease, a barrier to effective treatment, is generally caused by mutations in the enzyme that disrupt inhibitor binding but still allow for substrate processing. Structural studies with mutant, inactive enzyme, have provided detailed information regarding how the substrates bind to the protease yet avoid resistance mutations; insights obtained inform the development of next generation therapeutics. Although structures have been obtained of complexes between substrate peptide and inactivated (D25N) protease, thermodynamic studies of peptide binding have been challenging due to low affinity. Peptides that bind tighter to the inactivated protease than the natural substrates would be valuable …
Design And Synthesis Of Hiv-1 Protease Inhibitors Incorporating Oxazolidinones As P2/P2' Ligands In Pseudosymmetric Dipeptide Isosteres, G. S. Kiran Kumar Reddy, Akbar Ali, Madhavi Nalam, Saima Anjum, Hong Cao, Robin Nathans, Celia Schiffer, Tariq Rana
Design And Synthesis Of Hiv-1 Protease Inhibitors Incorporating Oxazolidinones As P2/P2' Ligands In Pseudosymmetric Dipeptide Isosteres, G. S. Kiran Kumar Reddy, Akbar Ali, Madhavi Nalam, Saima Anjum, Hong Cao, Robin Nathans, Celia Schiffer, Tariq Rana
Celia A. Schiffer
A series of novel HIV-1 protease inhibitors based on two pseudosymmetric dipeptide isosteres have been synthesized and evaluated. The inhibitors were designed by incorporating N-phenyloxazolidinone-5-carboxamides into the hydroxyethylene and (hydroxyethyl)hydrazine dipeptide isosteres as P2 and P2' ligands. Compounds with (S)-phenyloxazolidinones attached at a position proximal to the central hydroxyl group showed low nM inhibitory activities against wild-type HIV-1 protease. Selected compounds were further evaluated for their inhibitory activities against a panel of multidrug-resistant protease variants and for their antiviral potencies in MT-4 cells. The crystal structures of lopinavir (LPV) and two new inhibitors containing phenyloxazolidinone-based ligands in complex with wild-type …
Evaluating The Substrate-Envelope Hypothesis: Structural Analysis Of Novel Hiv-1 Protease Inhibitors Designed To Be Robust Against Drug Resistance, Madhavi Nalam, Akbar Ali, Michael Altman, G. S. Kiran Kumar Reddy, Sripriya Chellappan, Visvaldas Kairys, Aysegul Ozen, Hong Cao, Michael Gilson, Bruce Tidor, Tariq Rana, Celia Schiffer
Evaluating The Substrate-Envelope Hypothesis: Structural Analysis Of Novel Hiv-1 Protease Inhibitors Designed To Be Robust Against Drug Resistance, Madhavi Nalam, Akbar Ali, Michael Altman, G. S. Kiran Kumar Reddy, Sripriya Chellappan, Visvaldas Kairys, Aysegul Ozen, Hong Cao, Michael Gilson, Bruce Tidor, Tariq Rana, Celia Schiffer
Celia A. Schiffer
Drug resistance mutations in HIV-1 protease selectively alter inhibitor binding without significantly affecting substrate recognition and cleavage. This alteration in molecular recognition led us to develop the substrate-envelope hypothesis which predicts that HIV-1 protease inhibitors that fit within the overlapping consensus volume of the substrates are less likely to be susceptible to drug-resistant mutations, as a mutation impacting such inhibitors would simultaneously impact the processing of substrates. To evaluate this hypothesis, over 130 HIV-1 protease inhibitors were designed and synthesized using three different approaches with and without substrate-envelope constraints. A subset of 16 representative inhibitors with binding affinities to wild-type …
Combating Susceptibility To Drug Resistance: Lessons From Hiv-1 Protease, Nancy King, Moses Prabu-Jeyabalan, Ellen Nalivaika, Celia Schiffer
Combating Susceptibility To Drug Resistance: Lessons From Hiv-1 Protease, Nancy King, Moses Prabu-Jeyabalan, Ellen Nalivaika, Celia Schiffer
Celia A. Schiffer
Drug resistance is a major obstacle in modern medicine. However, resistance is rarely considered in drug development and may inadvertently be facilitated, as many designed inhibitors contact residues that can mutate to confer resistance, without significantly impairing function. Contemporary drug design often ignores the detailed atomic basis for function and primarily focuses on disrupting the target's activity, which is necessary but not sufficient for developing a robust drug. In this study, we examine the impact of drug-resistant mutations in HIV-1 protease on substrate recognition and demonstrate that most primary active site mutations do not extensively contact substrates, but are critical …
Decomposing The Energetic Impact Of Drug Resistant Mutations In Hiv-1 Protease On Binding Drv, Yufeng Cai, Celia Schiffer
Decomposing The Energetic Impact Of Drug Resistant Mutations In Hiv-1 Protease On Binding Drv, Yufeng Cai, Celia Schiffer
Celia A. Schiffer
Darunavir (DRV) is a high affinity (4.5x10(-12) M, DeltaG = -15.2 kcal/mol) HIV-1 protease inhibitor. Two drug-resistant protease variants FLAP+ (L10I, G48V, I54V, V82A) and ACT (V82T, I84V) decrease the binding affinity with DRV by 1.0 kcal/mol and 1.6 kcal/mol respectively. In this study the absolute and relative binding free energies of DRV with wild-type protease, FLAP+ and ACT were calculated with MM-PB/GBSA and thermodynamic integration methods, respectively. Free energy decomposition elucidated that the mutations conferred resistance by distorting the active site of HIV-1 protease so that the residues that lost binding free energy were not limited to the sites …