Structure and dynamics of the membrane attaching nitric oxide transporter nitrophorin 7 [version 1; peer review: 2 approved, 1 approved with reservations]

Nitrophorins represent a unique class of heme proteins that are able to perform the delicate transportation and release of the free-radical gaseous messenger nitric oxide (NO) in a pH-triggered manner. Besides its ability to bind to phospholipid membranes, the N-terminus contains an additional Leu-Pro-Gly stretch, which is a unique sequence trait, and the heme cavity is significantly altered with respect to other nitrophorins. These distinctive features encouraged us to solve the Xray crystallographic structures of NP7 at low and high pH and bound with different heme ligands (nitric oxide, histamine, imidazole). The overall fold of the lipocalin motif is well preserved in the different Xray structures and resembles the fold of other nitrophorins. However, a chain-like arrangement in the crystal lattice due to a number of head-to-tail electrostatic stabilizing interactions is found in NP7. Furthermore, the X-ray structures also reveal ligand-dependent changes in the orientation of the heme, as well as in specific interactions between the A-B and G-H loops, which are considered to be relevant for the biological function of nitrophorins. Fast and ultrafast laser triggered ligand rebinding experiments demonstrate the pH-dependent ligand migration within the cavities and the exit route. Finally, the topological distribution of pockets located around the heme as well as from inner cavities present at the rear of the protein provides a distinctive feature in NP7, so that while a loop gated exit mechanism to the solvent has been proposed for most Open Peer Review

[1]  Loriene Roy,et al.  What Is a Reference Source? , 2018, The Reference Librarian.

[2]  M. Knipp,et al.  Preparation of nitrophorin 7(Δ1-3) from Rhodnius prolixus without start-methionine using recombinant expression in Escherichia coli. , 2014, Analytical biochemistry.

[3]  Juan P. Bustamante,et al.  Ligand uptake modulation by internal water molecules and hydrophobic cavities in hemoglobins. , 2014, The journal of physical chemistry. B.

[4]  H. Ogata,et al.  Expression, purification, and solid-state NMR characterization of the membrane binding heme protein nitrophorin 7 in two electronic spin states. , 2013, Biochemistry.

[5]  F. J. Luque,et al.  Kinetics and computational studies of ligand migration in nitrophorin 7 and its Δ1-3 mutant. , 2013, Biochimica et biophysica acta.

[6]  F. J. Luque,et al.  Ligand migration through hemeprotein cavities: insights from laser flash photolysis and molecular dynamics simulations. , 2013, Physical chemistry chemical physics : PCCP.

[7]  Adrian E. Roitberg,et al.  pH-Dependent Conformational Changes in Proteins and Their Effect on Experimental pKas: The Case of Nitrophorin 4 , 2012, PLoS Comput. Biol..

[8]  Pier Remigio Salvi,et al.  Following Ligand Migration Pathways from Picoseconds to Milliseconds in Type II Truncated Hemoglobin from Thermobifida fusca , 2012, PloS one.

[9]  C. Viappiani,et al.  Heterogeneous kinetics of the carbon monoxide association and dissociation reaction to nitrophorin 4 and 7 coincide with structural heterogeneity of the gate-loop. , 2012, Journal of the American Chemical Society.

[10]  M. H. Pereira,et al.  Identification of the native N-terminus of the membrane attaching ferriheme protein nitrophorin 7 from Rhodnius prolixus. , 2012, Analytical biochemistry.

[11]  H. Ogata,et al.  Guanidine-ferroheme coordination in the mutant protein nitrophorin 4(L130R). , 2012, Angewandte Chemie.

[12]  R. Kosinsky,et al.  Insertion of Heme b into the Structure of the Cys34‐Carbamidomethylated Human Lipocalin α1‐Microglobulin: Formation of a [(Heme)2(α1‐Microglobulin)]3 Complex , 2012, Chembiochem : a European journal of chemical biology.

[13]  F. Javier Luque,et al.  MDpocket: open-source cavity detection and characterization on molecular dynamics trajectories , 2011, Bioinform..

[14]  Chunmao He,et al.  Reduction of the lipocalin type heme containing protein nitrophorin -- sensitivity of the fold-stabilizing cysteine disulfides toward routine heme-iron reduction. , 2011, Journal of inorganic biochemistry.

[15]  S. Neya,et al.  Breaking the proximal Fe(II)-N(His) bond in heme proteins through local structural tension: lessons from the heme b proteins nitrophorin 4, nitrophorin 7, and related site-directed mutant proteins. , 2011, Biochemistry.

[16]  Chunmao He,et al.  Nitrophorins: Nitrite disproportionation reaction and other novel functionalities of insect heme‐based nitric oxide transport proteins , 2011, IUBMB life.

[17]  A. Heller.  Electrochemistry and nitric oxide mass transport in cancer: why ingestion of sodium nitrite could be effective in treating vascularized tumors. , 2010, Physical chemistry chemical physics : PCCP.

[18]  H. Ogata,et al.  Formation of the complex of nitrite with the ferriheme b beta-barrel proteins nitrophorin 4 and nitrophorin 7. , 2010, Biochemistry.

[19]  Elspeth F. Garman,et al.  Radiation damage in macromolecular crystallography: what is it and why should we care? , 2010, Acta crystallographica. Section D, Biological crystallography.

[20]  M. Kubo,et al.  Ultrafast dynamics of diatomic ligand binding to nitrophorin 4. , 2010, Journal of the American Chemical Society.

[21]  F. Walker,et al.  1H and 13C NMR spectroscopic studies of the ferriheme resonances of three low-spin complexes of wild-type nitrophorin 2 and nitrophorin 2(V24E) as a function of pH , 2009, JBIC Journal of Biological Inorganic Chemistry.

[22]  T. Konuma,et al.  Structural dynamics and folding of beta-lactoglobulin probed by heteronuclear NMR. , 2009, Biochimica et biophysica acta.

[23]  K. Djinović-Carugo,et al.  Can soaked-in scavengers protect metalloprotein active sites from reduction during data collection? , 2009, Journal of synchrotron radiation.

[24]  A. Roitberg,et al.  pH-dependent mechanism of nitric oxide release in nitrophorins 2 and 4. , 2009, The journal of physical chemistry. B.

[25]  Marcelo A Martí,et al.  Molecular basis for the pH dependent structural transition of Nitrophorin 4. , 2009, The journal of physical chemistry. B.

[26]  Hongjun Zhang,et al.  A one-residue switch reverses the orientation of a heme b cofactor. Investigations of the ferriheme NO transporters nitrophorin 2 and 7 from the blood-feeding insect Rhodnius prolixus. , 2009, Biochemistry.

[27]  A. Roitberg,et al.  Bond or cage effect: how nitrophorins transport and release nitric oxide. , 2008, Journal of the American Chemical Society.

[28]  M. Shokhirev,et al.  Spectroscopic and functional characterization of nitrophorin 7 from the blood-feeding insect Rhodnius prolixus reveals an important role of its isoform-specific N-terminus for proper protein function. , 2007, Biochemistry.

[29]  Giulio Cerullo,et al.  High-time-resolution pump-probe system with broadband detection for the study of time-domain vibrational dynamics. , 2007, The Review of scientific instruments.

[30]  D. Kondrashov,et al.  Nonequilibrium dynamics simulations of nitric oxide release: comparative study of nitrophorin and myoglobin. , 2007, The journal of physical chemistry. B.

[31]  S. Mangani,et al.  Crystallization of soluble proteins in vapor diffusion for x-ray crystallography , 2007, Nature Protocols.

[32]  F. Walker,et al.  Overexpression in Escherichia coli and functional reconstitution of the liposome binding ferriheme protein nitrophorin 7 from the bloodsucking bug Rhodnius prolixus. , 2007, Protein expression and purification.

[33]  Kevin M. Smith,et al.  Assignment of the ferriheme resonances of the low-spin complexes of nitrophorins 1 and 4 by (1)H and (13)C NMR spectroscopy: comparison to structural data obtained from X-ray crystallography. , 2007, Inorganic chemistry.

[34]  F. Spyrakis,et al.  The reactivity with CO of AHb1 and AHb2 from Arabidopsis thaliana is controlled by the distal HisE7 and internal hydrophobic cavities. , 2007, Journal of the American Chemical Society.

[35]  N. Lehnert,et al.  Spectroscopic properties and electronic structure of five- and six-coordinate iron(II) porphyrin NO complexes: Effect of the axial N-donor ligand. , 2006, Inorganic chemistry.

[36]  C. Renner,et al.  Arginine- and lysine-specific polymers for protein recognition and immobilization. , 2006, Journal of the American Chemical Society.

[37]  Laxmikant V. Kalé,et al.  Scalable molecular dynamics with NAMD , 2005, J. Comput. Chem..

[38]  A. Mozzarelli,et al.  Determination of microscopic rate constants for CO binding and migration in myoglobin encapsulated in silica gels. , 2005, The journal of physical chemistry. B.

[39]  G. Keserü,et al.  Protonation state of Asp30 exerts crucial influence over surface loop rearrangements responsible for NO release in nitrophorin 4 , 2005, FEBS letters.

[40]  J. Larsson,et al.  The lipocalin alpha(1)-microglobulin binds heme in different species. , 2004, Archives of biochemistry and biophysics.

[41]  D. Kondrashov,et al.  Protein functional cycle viewed at atomic resolution: conformational change and mobility in nitrophorin 4 as a function of pH and NO binding. , 2004, Biochemistry.

[42]  J. Andersen,et al.  Recognition of anionic phospholipid membranes by an antihemostatic protein from a blood-feeding insect. , 2004, Biochemistry.

[43]  J. Andersen,et al.  Role of binding site loops in controlling nitric oxide release: structure and kinetics of mutant forms of nitrophorin 4. , 2004, Biochemistry.

[44]  A. Mozzarelli,et al.  CO rebinding kinetics to myoglobin- and R-state-hemoglobin-doped silica gels in the presence of glycerol , 2004 .

[45]  F. Walker,et al.  Axial ligand complexes of the Rhodnius nitrophorins: reduction potentials, binding constants, EPR spectra, and structures of the 4-iodopyrazole and imidazole complexes of NP4 , 2004, JBIC Journal of Biological Inorganic Chemistry.

[46]  J. Mccleverty.  Chemistry of nitric oxide relevant to biology. , 2004, Chemical reviews.

[47]  W. Scheidt,et al.  Five- to six-coordination in (nitrosyl)iron(II) porphyrinates: effects of binding the sixth ligand. , 2003, Inorganic chemistry.

[48]  D R Flower,et al.  The lipocalin protein family: structural and sequence overview. , 2000, Biochimica et biophysica acta.

[49]  J. Andersen,et al.  The Crystal Structure of Nitrophorin 2 , 2000, The Journal of Biological Chemistry.

[50]  J. Andersen,et al.  Nitric oxide binding to nitrophorin 4 induces complete distal pocket burial , 2000, Nature Structural Biology.

[51]  Alexander D. MacKerell,et al.  All-atom empirical potential for molecular modeling and dynamics studies of proteins. , 1998, The journal of physical chemistry. B.

[52]  Wolfgang Kabsch,et al.  Automatic processing of rotation diffraction data from crystals of initially unknown symmetry and cell constants , 1993 .

[53]  M. Lehane,et al.  The biology of blood-sucking in insects , 1991 .

[54]  W. L. Jorgensen,et al.  Comparison of simple potential functions for simulating liquid water , 1983 .

[55]  H. Ogata,et al.  Crystallization and preliminary X-ray crystallographic analysis of the membrane-binding haemprotein nitrophorin 7 from Rhodnius prolixus. , 2012, Acta crystallographica. Section F, Structural biology and crystallization communications.

[56]  Klaus Schulten,et al.  Finding gas migration pathways in proteins using implicit ligand sampling. , 2008, Methods in enzymology.

[57]  C. Schulze-Briese,et al.  Cryoradiolytic reduction of crystalline heme proteins: analysis by UV-Vis spectroscopy and X-ray crystallography. , 2007, Journal of synchrotron radiation.

[58]  F. Walker.  Nitric oxide interaction with insect nitrophorins and thoughts on the electron configuration of the {FeNO}6 complex. , 2005, Journal of inorganic biochemistry.

[59]  P. Afonine,et al.  research papers Acta Crystallographica Section D Biological , 2003 .

[60]  A. Romanha,et al.  Identification of morphologically similar Rhodnius species (Hemiptera: Reduviidae: Triatominae) by electrophoresis of salivary heme proteins. , 2000, The American journal of tropical medicine and hygiene.

[61]  J. Andersen,et al.  Nitric Oxide Binding to the Ferri- and Ferroheme States of Nitrophorin 1, a Reversible NO-Binding Heme Protein from the Saliva of the Blood-Sucking Insect, Rhodnius prolixus , 1999 .