Biomedical subjects

K H Mayo

Publications and source records attributed to K H Mayo.

At least 73 records · Page 4Linked to original sources

PubMed1994

Subunit association and structural analysis of platelet basic protein and related proteins investigated by 1H NMR spectroscopy and circular dichroism.

Platelet basic protein (PBP) (94 residues) is naturally processed via N-terminal cleavage to yield connective tissue activating peptide-III (85 residues), beta-thromboglobulin (81 residues), and neutrophil activating peptide-2 (70 residues). Chemical cross-linking and gel filtration data indicate that each homolog can form dimers and tetramers. Subunit association equilibria for dimer (KD) and tetramer (KT) formation have been derived for each species from 1H NMR (600 MHz) spectral analysis of slowly exchanging (NMR time scale) monomer- dimer-tetramer aggregation state populations. In general, raising the pH from about pH 3.5 to pH 6 increases KD by two to three orders in magnitude and decreases KT by some 50-fold. Ionic strength effects also suggest that intersubunit electrostatic interactions are critical to subunit association. Subunit stabilization can be ranked proportional to N-terminal chain length: platelet basic protein > connective tissue activating peptide-III > beta-thromboglobulin > neutrophil activating peptide-2. Under more physiologic conditions, PBP family monomers are favored at normal cytokine protein concentrations and may form the biologically active state. CD and NMR data indicate conservation of alpha-helix and anti-parallel beta-sheet structure among PBP-related species and support the idea that the extended N terminus folds over and masks the neutrophil activation domain and is part of the intersubunit binding domain.

Amino Acid Sequence↗

PubMed1994

Rhenium-labeled somatostatin analog RC-160. 1H NMR and computer modeling conformational analysis.

The solution conformations of RC-160, cyclic D-Phe1-Cys2-Tyr3-D-Trp4-Lys5-Val6-Cys7+ ++-Trp8-NH2, an analog of the tumor antiproliferatory neuropeptide somatostatin, and RC-160 labeled with rhenium (Re-RC-160), have been determined by using two-dimensional 1H NMR spectroscopy (600 MHz) and restrained molecular dynamics simulations. Re-RC-160 yields the same average solution conformation as does the apo form with an antiparallel beta-sheet fold and a type II' beta-turn centered at D-Trp4-Lys5. These results indicate that the spatial topography of the side chains essential for somatostatin receptor binding is maintained in Re-RC-160.

Amino Acid Sequence↗

PubMed1993

Tri- and diglycine backbone rotational dynamics investigated by 13C NMR multiplet relaxation and molecular dynamics simulations.

Backbone motional dynamics in tri- and diglycine have been investigated by using 13C NMR multiplet relaxation spectroscopy. Dipolar auto- and cross-correlation times were determined as a function of pH, ionic strength, and temperature. Molecular dynamics simulations and phi,psi bond rotation energy profiles were calculated for insight into the physical nature of backbone rotations that could contribute to 13C relaxation. Various motional models were used to fit the experimental data. For internal glycine G2 in triglycine, restricted and unrestricted rotational diffusion models both underestimate internal correlation times, although they do agree that the axis of fastest internal rotation is directed closely along the C alpha-C bond. For di- and triglycine, significant pH dependencies in cross-correlation times for C-terminal glycines, and more so for those of N-terminal glycines, indicate the importance of the ionization state in internal mobility of terminal backbone positions. For terminal glycines, rotational jump models which allow for diffusive-like fluctuations within minima best explain the experimental data. phi,psi rotational fluctuation amplitudes and internal rotational energy barriers derived from the temperature dependence of 13C relaxation parameters, which range from 3 to 5 kcal/mol, agree well with those values calculated in rotational energy profiles.

Amino Acid Sequence↗

PubMed1993

13C nuclear magnetic resonance relaxation-derived psi, phi bond rotational energy barriers and rotational restrictions for glycine 13C alpha-methylenes in a GXX-repeat hexadecapeptide.

Spin-lattice relaxation of 13C multiplet spectra and [1H]-13C nuclear Overhauser enhancement (NOE) coefficients of selectively 13C-enriched glycines in a collagen GXX-repeat motif hexadecapeptide, G1VKG4DKG7NPG10WPG13APY, has been investigated. Data have been collected at two 13C Larmor frequencies (90 and 150 MHz) over the temperature range from 5 to 70 degrees C. Relaxation data indicate that the most restricted internal rotations are at G7 and G10. Mobility of other glycine residues can be arranged in the order G4, G13, and G1. G1 glycine shows that least change in motional anisotropy with temperature. Several motional models have been used to explain the experimental data. While any one model is not completely satisfactory in describing all experimental parameters, only the model of restricted internal diffusion yields the observed positive sign for the cross-correlated spectral densities. Energetic and angular limits of psi, phi bond rotational motions derived from relaxation data and the restricted diffusion model are in good agreement with those calculated as Ramachandran potentional energy profiles. G1 rotational energy barriers for overall tumbling and internal rotation are approximately equal, suggesting strong interaction between the N-terminus and water. Internal rotational parameters for GV and GG dipeptides confirm this view. Nonterminal glycine internal motions are apparently less dependent on water-peptide interactions.

Amino Acid Sequence↗

PubMed1993

Alcohol-induced protein folding transitions in platelet factor 4: the O-state.

Platelet factor 4 (PF4) (7800 daltons) is an anti-parallel beta-sheet, alpha/beta class protein whose tertiary structure is stabilized by the presence of two disulfide bonds. Titration of PF4 with 2-propanol or similar low molecular weight, aliphatic alcohols induces reversible protein folding transitions which are observed to be in slow exchange on the 600-MHz 1H NMR time scale. Line fitting of resolved resonances assigned to ring protons of Y60, H35, H23, and alpha H of K50 in native and alcohol-induced states (O-states) allows derivation of folding equilibrium constants and exchange kinetics. Folding exchange rates vary between 5 and 100 s-1 on going from 9.8 to 3.3 M2-propanol. Simple linear extrapolation to 0 M2-propanol yields an O-state to N-state exchange rate of about 500 s-1, i.e., millisecond time scale. At relatively high 2-propanol concentration (>9.5 M), where the O-state is predominant (>90%), NMR spectra suggest a more "unfolded" structure, while CD data indicate the preservation of considerable secondary structure. Increasing 2-propanol from 3.3 to 9.8 M, however, shifts the CD-derived fractional compositions significantly, with overall beta-structure decreasing by about 20% and alpha-helix composition increasing by about 25%. Alcohol-jump experiments, which identify O-state long-lived NHs in the NMR spectrum of native PF4, indicate folding transition reversibility and conservation of about 15 long-lived NHs in native and O-states. Most of these NHs are assigned to residues in anti-parallel beta-sheet structure. Of these 15 NHs, H/D exchange rates, although variably reduced in the O-state, are generally still long-lived compared with random coil H/D exchange. Overall, the PF4 O-state is a stable intermediate with an apparently more highly fluctuating anti-parallel beta-sheet structure and a more stabilized C-terminal alpha-helix.

1-Propanol↗

PubMed1993

Structural features of fibronectin synthetic peptide FN-C/H II, responsible for cell adhesion, neurite extension, and heparan sulfate binding.

FN-C/H II (KNNQKSEPLIGRKKT), a heparin-binding peptide derived from the COOH-terminal heparin-binding domain of fibronectin, mediates cell adhesion for a variety of cell types and promotes neurite outgrowth. By systematic amino acid substitution of synthetic peptide analogues of FN-C/H II, the basic structural features necessary for activity have been identified in the COOH-terminal residues LIGRKK. This biologically "active" sequence has been located in several other heparin/heparan sulfate-binding proteins and may represent a potential binding motif for sulfated polyanions. NMR structural studies indicate that the COOH-terminal segment of FN-C/H II displays significant multiple-turn or helix-like character suggesting that the RKK sequence may lie on the same surface of the protein, as opposed to alternating in an extended chain motif.

Amino Acid Sequence↗

PubMed1993

Effect of low molecular weight aliphatic alcohols and related compounds on platelet factor 4 subunit association.

Titration of platelet factor 4 (PF4) with increasing concentrations of various low molecular weight aliphatic alcohols disrupts dimer and tetramer quaternary structure while preserving tertiary structural integrity. Normally observed slow subunit exchange (1H NMR time scale) is shifted into the fast chemical exchange regime. The order of effectiveness is butanol > 2-propanol > propanol > ethanol > methanol. Fluorination of ethanol (trifluoroethanol) and 2-propanol (hexafluoro-2-propanol) increases effectiveness by approximately 2 orders of magnitude. Oxidation to 2-propanone or trifluoro-2-propanone produces a less effective compound, suggesting a mechanistic role for the hydroxyl group. Increasing the number of hydroxyls to three, as in glycerol, however, is relatively ineffective in disrupting aggregate states or modifying subunit exchange rates. While 19F NMR studies indicate that these alcohols specifically interact with PF4, binding alone can not explain their mechanism of action. Dimethyl sulfoxide, structurally similar to 2-propanol, disrupts PF4 aggregation by direct binding, but does not shift subunit exchange kinetics into the NMR fast-exchange regime. Although not fully understood, the effectiveness of these compounds appears to be related to the colligative properties of the solution.

Alcohols↗

PubMed1993

Effects of molecular association on structure and dynamics of a collagenous peptide.

Peptide GVKGDKGNPGWPGAPY from the triple-helix domain of type IV collagen aggregates in solution at a critical aggregation concentration of 18 mM. This molecular self-association process is investigated by 1H- and 13C-nmr spectroscopy. As a function of increasing peptide concentration, selective 1H resonances are cooperatively chemically shifted by up to 0.04 ppm to apparently saturable values at high concentration. Pulsed field gradient nmr was used to derive translation diffusion constants that, as the peptide concentration is increased, also cooperatively and monotonically decrease to an apparent limiting value. An average number of 6 monomer units per aggregate have been estimated from diffusion constant and 13C relaxation data. Comparative 1H nuclear Overhauser effect spectroscopy (NOESY) spectra accumulated at high and low peptide concentrations suggest that average internuclear distances are decreased as a result of peptide association. 13C-nmr multiple spin-lattice relaxation and 13C-[1H] NOE effects on 13C-enriched glycine methylene positions in the peptide demonstrate that overall molecular tumbling and backbone internal motions are attenuated in the aggregate state. Lowering the solution pD from pD 6 to pD 2 disrupts the aggregate state, suggesting a role for electrostatic interactions in the association process. Based on thermodynamic considerations, hydrophobic interactions also probably act to stabilize the aggregate state. These data are discussed in terms of an nmr/NOE constrained computer-modeled structure of the peptide.

Amino Acid Sequence↗

PubMed1993

pH sensitivity of epidermal growth factor receptor complexes.

The association/dissociation binding kinetics of 125I-labeled mouse epidermal growth factor (EGF) to receptors on human fibroblast cells in monolayer culture have been measured at 4 degrees C as a function of extracellular pH from pH 5-9. At pH 8, steady-state total binding is maximal. As pH is lowered to 6.5, total binding monotonically decreases dramatically. It changes further only slightly between pH 6.5 and 5 to about 20% of the maximum binding value. Scatchard binding plots at pH 7.5 and above show the commonly observed concave-upward, non-linear curve; as pH is lowered, this plot becomes much more linear, indicating that the "high affinity" bound receptor population is greatly diminished. Application of our ternary complex binding model [Mayo et al., J Biol Chem 264:17838-17844, 1989], which hypothesizes complexation of the EGF-bound receptor with a cell surface interaction molecule, indicates that pH may have some direct effects on ternary complex formation, but the major effect is on EGF-receptor dissociation.

ErbB Receptors↗

PubMed1992

Molten globule monomer to condensed dimer: role of disulfide bonds in platelet factor-4 folding and subunit association.

Platelet factor 4 (PF4) exhibits high affinity for heparin and exists as a tetramer in solution under physiologic conditions. Reduction of the two disulfide bridges in PF4 increases the protein's dissociation constant for heparin approximately 20-fold and shifts the highest apparent aggregation state from tetramer to dimer as evidenced by gel filtration, chemical cross-linking, and 1H-NMR studies. 1H-NMR spectra of reduced PF4 monomers generally show narrower, less dispersed, upfield-shifted NH and alpha H resonances, suggesting the presence of an unfolded monomer state. Reduced PF4 monomer folding, however, is evidenced by the presence of about 12 relatively long-lived backbone NHs and by CD spectra that indicate conservation of overall secondary structure. These data suggest the presence of a molten globule-type state. Urea denaturation shifts this apparent molten globule to a fully unfolded state characterized by more random coil-like resonance shifts. The reduced PF4 dimer state yields NMR and CD data consistent with preservation of tertiary structural folds found for the native species. In this regard, the reduced PF4 folding transition is thermodynamically linked with dimer formation which stabilizes tertiary structure. Monomer-dimer association equilibria for reduced PF4 essentially follow the same pH and salt titration trends as reported previously for native PF4 dimers [Mayo, K. H., & Chen, M. J. (1989) Biochemistry 28, 9469-9478], indicating that that dimer interface is generally conserved in the absence of disulfide constraints. Reduced PF4 tetramers are not apparent under any conditions investigated, suggesting that disulfides are necessary for efficient antiparallel beta-sheet alignment between dimer pairs.

Chromatography, Gel↗

PubMed1991

1H NMR and CD secondary structure analysis of cell adhesion promoting peptide F-9 from laminin.

A heparin binding, cell adhesion promoting domain, termed peptide F-9, from the B1 chain of human laminin, residues 641 to 660, i.e. RYVVLPRPVCFEKGMNYTVR, has been investigated by 1H NMR (500 MHz) spectroscopy and CD spectropolarimetry. While small linear peptides in water solution normally exist in a number of fluctuating conformational states, CD data analysis of peptide F9 indicates the existence of some preferred average structural populations consisting of about 30% beta-sheet, 22% beta-turn, and 6% alpha-helix. NMR structural analysis supports this observation and indicates specific sequences of preferred structural populations. Evidence for these is indicated by the presence of dNN nuclear Overhauser effect (NOE) populations and attenuated or absent d alpha N NOEs at short mixing times (0.1 s), 3J alpha N coupling constants of 5 and 10 Hz, and chemical shifts significantly removed from random coil positions. The NH2-terminal VVL sequence primarily exists in an extended chain conformation by virtue of large d alpha N NOEs and 9-10 Hz 3J alpha N coupling constants. Residues C10-N16 have turn-like or helix character with a run of dNN and d beta N NOEs and attenuated d alpha N NOEs. These midchain reversals include the lysine and asparagine residues proposed to be involved in heparin binding and N-glycosylation, respectively, to laminin peptide F-9.

Amino Acid Sequence↗

PubMed1991

Cell adhesion promoting peptide GVKGDKGNPGWPGAP from the collagen type IV triple helix: cis/trans proline-induced multiple 1H NMR conformations and evidence for a KG/PG multiple turn repeat motif in the all-trans proline state.

Peptide GVKGDKGNPGWPGAPY (called peptide IV-H1), derived from the protein sequence of human collagen type IV, triple-helix domain residues 1263-1277, represents an RGD-independent, cell-specific, adhesion, spreading, and motility promoting domain in type IV collagen. In this study, peptide IV-H1 has been investigated by 1H NMR (500 MHz) spectroscopy. Cis-trans proline isomerization at each of the three proline residues gives rise to a number of slowly exchanging (500-MHz NMR time scale) conformation states. At least five such states are observed, for example, for the well-resolved A14 beta H3 group, and K3, which is six residues sequentially removed from the nearest proline, i.e., P9, shows two sets. The presence of more than two sets of resonances for residues sequentially proximal to a proline, e.g., A14-cis-P15 and A14-trans-P15, and more than one set for a residue sequentially well-removed from a proline, e.g., K3, indicates long range conformation interactions and the presence of preferred structure in this short linear peptide. Many resonances belonging to these multiple species have been assigned by using mono-proline-substituted analogues. Conformational (isomer) state-specific 2D 1H NMR assignments for the combination of cis and trans proline states have been made via analysis of COSY-type, HOHAHA, and NOESY spectra. Peptide IV-H1 in the all-trans proline state ttt exists in relatively well-defined conformation populations showing numerous short- and long-range NOEs and long-lived backbone amide protons and reduced backbone NH temperature coefficients, suggesting hydrogen-bonding, and structurally informative 3J alpha N coupling constants. The NMR data indicate significant beta-turn populations centered at K3-G4, K5-G6, P9-G10, and P12-G13, and a C-terminal gamma-turn within the A14-P15-Y16 sequence. These NMR data are supported by circular dichroic studies which indicate the presence of 52% beta-turn, 10% helix, and 38% random coil structural populations. Since equally spaced KG and PG residues are found on both sides of peptide IV-H1 in the native collagen type IV sequence, this multiple turn repeat motif may continue through a longer segment of the protein. Synthetic peptide IV-H1 overlapping sequence "walk throughs" indicate that the primary biological activity is localized in the GNPGWPGAP double beta-turn domain, which contains the backbone constraining proline residues. This proline-domain conformation may suggest a collagen type IV receptor-specific, metastatic cell adhesion promoting binding domain.

Amino Acid Sequence↗

PubMed1991

Human platelet factor 4 subunit association/dissociation thermodynamics and kinetics.

Platelet factor 4 (PF4) monomers (7800 daltons) form dimers and tetramers in varying molar ratios under certain solution conditions [Mayo, K. H., & Chen, M. J. (1989) Biochemistry 28, 9469]. The presence of a simplified aromatic region (one Tyr and two His) and resolved monomer, dimer, and tetramer Y60 3,5 ring proton resonances makes study of PF4 aggregate association/dissociation thermodynamics and kinetics possible. PF4 protein subunit association/dissociation equilibrium thermodynamic parameters have been derived by 1H NMR (500MHz) resonance line-fitting analysis of steady-state Y60 3,5 ring proton resonance monomer-dimer-tetramer populations as a function of temperature from 10 to 40 degrees C. Below 10 degrees C and above 40 degrees C, resonance broadening and overlap severely impaired analysis. Enthalpic and entropic contributions to dimer association Gibb's free energy [-5.1 kcal/mol (30 degrees C)] are +2.5 +/- 1 kcal/mol and +26 +/- 7 eu, respectively, and for tetramer association Gibb's free energy [-5.7 kcal/mol (30 degrees C)], they are -7.5 +/- 1 kcal/mol and -7 +/- 3 eu, respectively. These thermodynamic parameters are consistent with low dielectric medium electrostatic/hydrophobic interactions governing dimer formation and hydrogen bonding governing tetramer formation. Association/dissociation kinetic parameters, i.e., steady-state jump rates, have been derived from exchange-induced line-width increases and from 1H NMR (500 MHz) saturation-transfer and spin-lattice (Tl) relaxation experiments. From dissociation jump rates and equilibrium constants, association rate constants were estimated. For dimer and tetramer equilibria at 30 degrees C, unimolecular dissociation rate constants are 35 +/- 10 s-1 for dimer dissociation and 6 +/- 2 s-1 for tetramer dissociation.(ABSTRACT TRUNCATED AT 250 WORDS)

Humans↗

PubMed1991

Low-affinity platelet factor 4 1H NMR derived aggregate equilibria indicate a physiologic preference for monomers over dimers and tetramers.

Low-affinity platelet factor 4 (LA-PF4), unlike another related, sequentially homologous (about 50%) platelet-specific protein, platelet factor 4 (PF4), is an active mitogenic and chemotactic agent. PF4 exhibits a high binding affinity for heparin, while LA-PF4 does not. Both PF4 and LA-PF4 can exist in dimer and tetramer aggregate states. Equilibrium constants for PF4 aggregation have recently been estimated from fractional populations derived from proton nuclear magnetic resonance (NMR) integrals assigned to resonances in monomer, dimer, and tetramer states [Mayo & Chen (1989) Biochemistry 28, 9469]. On a 500-MHz NMR time scale, relatively slow exchange among LA-PF4 aggregate species has also allowed Tyr 15 ring proton resonances to be assigned for monomer, dimer, and tetramer states in LA-PF4. As a function of pH and ionic strength, equilibrium association constants for LA-PF4 dimer (KD) and tetramer (KT) formation have been estimated from Tyr 15 ring proton resonance integrals. At low ionic strength, KD reaches a minimum value of 12 M-1 at pH 3 where KT is at its maximum value of 1.6 x 10(5) M-1. At pH 4.1, KD and KT have the same value, 1.1 x 10(3) M-1, which is the minimum value for KT. KD plateaus off to its maximum value of 2.2 x 10(4) M-1 by pH 5.5. These values are significantly lower than those for PF4. Analysis of the pH dependence of KD and KT suggests that electrostatic interactions probably among Glu/Asp and Lys/Arg side chains form the predominant force in the monomer-monomer binding process, i.e., KD, while like-charge repulsion due to proximal, intersubunit Glu/Asp residues decreases KT as the pH is raised. At pH 7 and low ionic strength, the dimer state is highly favored over the tetramer state. Elevating the solvent ionic strength at pH 7 destabilizes the dimer state. Under these more physiologic conditions, i.e., pH 7 and 0.1-0.2 M NaCl, LA-PF4 monomers are highly favored over dimers and tetramers. For PF4 under similar solvent conditions, tetramers predominate. Differences in biological activities between these homologous platelet-specific proteins may be the result, at least in part, of differing aggregation properties. The biologically active state for PF4 is tetramer, while for LA-PF4 it is monomer. Quaternary structure may, therefore, account for strong heparin binding in PF4, most likely by presenting a more favorable structural matrix for effective glycosaminoglycan interactions.

Amino Acid Sequence↗

PubMed1991

Attitude variables of dentofacial deformity patients: demographic characteristics and associations.

For patients to obtain satisfaction from surgical orthodontic treatment, a concordance needs to exist between the patients' concerns and expectations and the clinician's outcome measures of success. In this study, 231 patients 16 years or older were analyzed to define attributes of treatment that relate to patient satisfaction with outcome. The most prevalent patient concerns were related to facial and dental esthetics, and the effect of the dentofacial deformity on the quality of life was associated with a significantly higher motivation for surgical treatment.

Adolescent↗

PubMed1990

Human platelet factor 4 and its C-terminal peptides: heparin binding and clearance from the circulation.

Human platelet factor 4 (PF4), a high affinity heparin binding protein, is released from stimulated platelets and stored at vascular sites, predominantly in liver, from where it can be brought back into circulation by heparin. We attempted to define structural requirements for PF4 binding to heparin and for the pattern of its clearance from the circulation. Intact PF4 bound strongly to heparin agarose and was eluted at 1.4 M NaCl, while reduced PF4 and PF4 C-terminal peptides PF4 (47-70) and PF4 (58-70) bound weakly and were eluted at 0.2-0.5 M NaCl. 125I-radiolabeled intact PF4, reduced PF4 and C-terminal PF4 peptides injected into rabbits were cleared from the circulation in a biphasic pattern with components having half-life time of 1-2 min and 20-140 min. Heparin eliminated the fast component of PF4 clearance, but it did not affect clearance of reduced PF4 or C-terminal PF4 peptides. In contrast to reduced PF4 and PF4 (47-70), intact PF4 that accumulated in the liver and spleen, was displaced by heparin into circulating blood. In conclusion, specific binding sites and native conformation of the molecule are critical for high affinity PF4 binding to insolubilized heparin and for a pattern of PF4 clearance from the circulation in the presence of heparin.

Animals↗

PubMed1990

Proton magnetic resonance spectroscopic analysis of diadenosine 5',5"'-polyphosphates.

Certain diadenosine 5',5"'-polyphosphates are potent inhibitors of ADP stimulated platelet aggregation, acting possibly via competitive ADP-receptor binding. 1H NMR studies of a series of such compounds where the number of phosphate groups between adenosine groups was varied from 2 to 6 were performed to analyze possible preferred solution conformations and to define structure-activity relations. Relative to mononucleotides ADP and ATP, chemical shifts of adenosine proton resonances in diadenosine polyphosphate analogs are upfield shifted suggesting base stacking. This effect is greatest for AP2A and AP3A. Coupling constants of ribose ring proton resonances support the idea of an anti-base-ribose ring conformation, and 3JH5'-P values suggest a preferred gauche H-C-O-P structure. In all cases, NMR parameters for AP2A are near-limiting values for a static base stacked conformation. Increasing the number of phosphate groups between adenosine moieties tends to weaken this interaction.

Adenine Nucleotides↗

PubMed1990

Divalent cation binding to reduced and octanoyl acyl-carrier protein.

Mn(II) EPR binding studies with reduced acyl-carrier protein (ACP-SH) strongly suggest the presence of two relatively high-affinity manganese-binding sites (average Kd/site approximately 80 microM) at physiological pH. Lowering the pH or titrating with sodium chloride reduces the average number of bound divalent cations and decreases the binding affinity. This is consistent with the idea that anionic ligand(s), e.g. the carboxylate of glutamic or aspartic acid, on the protein are involved in manganese ion coordination. At pH values above 8.0, binding affinity is also reduced, whereas the average number of bound metal ions increases to about five at pH 8.5. By interacting weakly with divalent cations (average Kd/site approximately 1 mM), octanoyl acyl-carrier protein (OcoACP) exhibits dramatically different metal-ion-binding properties compared to ACP-SH. Calcium and magnesium can compete in either ACP species for manganese binding. Photochemically-induced dynamic nuclear polarisation 1H-NMR experiments strongly suggest that ACP-SH and OcoACP undergo at pH-induced conformational change between pH 5.5 and pH 7.0, and that divalent cations stabilize the protein against such pH-induced structural perturbations.

Acyl Carrier Protein↗