Glutamic Acid Side Chain Research Articles

Synthesis and Macrodomain Binding of Gln-carba-ADPr-peptide.

Mono-ADP-ribosylation is a dynamic post-translational modification (PTM) with important roles in cell signalling. This modification occurs on a wide variety of amino acids, and one of the canonical modification sites within proteins is the side chain of glutamic acid. Given the transient nature of this modification (acylal linkage) and the high sensitivity of ADP-ribosylated glutamic acid, stabilized isosteres are required for structural and biochemical studies. Here, we report the synthesis of a mimic of ADP-ribosylated peptide derived from histone H2B that contains carba-ADP-ribosylated glutamine as a potential mimic for Glu-ADPr. We synthesized a cyclopentitol-ribofuranosyl derivative of 5'-phosphoribosylated Fmoc-glutamine and used this in the solid-phase synthesis of the carba-ADPr-peptide mimicking the ADP-ribosylated N-terminal tail of histone H2B. Binding studies with isothermal calorimetry demonstrate that the macrodomains of human MacroD2 and TARG1 bind to carba-ADPr-peptide in the same way as ADPr-peptides containing the native ADP-riboside moiety connected to the side chain of glutamine in the same peptide sequence.

Controlling Helical Asymmetry in Supramolecular Copolymers by In Situ Chemical Modification.

Amplification of asymmetry in complex molecular systems results from a delicate interplay of chiral supramolecular structures and their chemical reactivity. In this work, we show how the helicity of supramolecular assemblies can be controlled by performing a non-stereoselective methylation reaction on comonomers. By methylating chiral glutamic acid side chains in benzene-1,3,5-tricarboxamide (BTA) derivatives to form methyl esters, the assembly properties are modulated. As reacted comonomers, the methyl ester-BTAs induce a stronger bias in the screw-sense of helical fibers predominantly composed of stacked achiral alkyl-BTA monomers. Hence, applying the in situ methylation in a system with the glutamic acid-BTA comonomer induces asymmetry amplification. Moreover, mixing small quantities of enantiomers of glutamic acid-BTA and glutamate methyl ester-BTA in the presence of the achiral alkyl-BTAs leads to deracemization and inversion of the helical structures in solution via the in situ reaction toward a thermodynamic equilibrium. Theoretical modeling suggests that the observed effects are caused by enhanced comonomer interactions after the chemical modification. Our presented methodology enables on-demand control over asymmetry in ordered functional supramolecular materials.

Phosphatidyl serine preconditions the A2AAR for G-protein coupling via a conserved motif on TM6 and TM7.

19F NMR of A2AAR in nanodiscs of defined composition indicates that negative charge modulates activation of the receptor. The fully active state is obtained either when bound to G-protein, or without G-protein but in a membrane environment containing negatively charged headgroups. Mutagenesis indicates that a trio of residues on the intracellular ends of TM6 and TM7 are responsible for the lipid-dependent activity. In order to determine the mechanism, a series of molecular dynamics simulations were performed. Comparison of simulations of the inactive and the active, G-protein bound state shows that a glutamic acid on the helix of the G-protein coordinates interactions between these three residues. This suggested a hypothesis in which a negatively charged headgroup can stand-in for the glutamic acid side chain, coordinating the ends of TM6 and TM7 and preconditioning the receptor for G-protein coupling. To test this idea, simulations of the fully active state were performed with and without phosphatidyl serine headgroups, but without the G-protein. Consistent with our hypothesis, the TM6 -- TM7 interactions are similar to the G-protein coupled state in the presence of PS, but similar to the inactive state in the absence of PS. Moreover, the interaction of a PS headgroup with a key residue on TM6 recapitulates the same interaction with the glutamic acid, but only when the receptor is in the active state.

Metal coordination to solute binding proteins - exciting chemistry with potential biological meaning.

Zn(II) is essential for bacterial survival and virulence. In host cells, its abundance is extremely limited, thus, bacteria have evolved transport mechanisms that enable them to take up this essential metal nutrient. Paracoccus denitrificans encodes two solute binding proteins (SBPs) - ZnuA and AztC, which are responsible for zinc acquisition from the host cells. We focus on understanding the interactions of Zn(II) and Ni(II) (zinc's potential competitor, which is a biologically relevant metal ion essential for various bacterial enzymes) with the extracellular ZnuA and AztC's loops from P. denitrificans that are expected to be possible Zn(II) binding sites. In the case of Zn(II) complexes with ZnuA outercellular loop regions, the numerous histidines act as anchoring donors, forming complexes with up to four coordinated His residues, while in the AztC region, three imidazole nitrogens and one water molecule are involved in Zn(II) binding. In Zn(II) complexes with ZnuA His-rich loop regions, so-called polymorphic binding sites are observed. The large number of available imidazoles and carboxylic side chains also strongly affects the structure of Ni(II) complexes; the more histidines in the studied peptide, the higher the affinity to bind Ni(II) and the higher the pH value at which amide nitrogens start to participate in Ni(II) binding. Additionally, for Ni(II)-ZnuA complexes, a more rare octahedral geometry is observed and such complexes are more stable than the corresponding Zn(II) ones, in contrast to what was observed in the AztC region, suggesting that the numerous histidyl and glutamic acid side chains are more tempting for Ni(II) than for Zn(II).The general strong affinity of Zn(II)-zincophore complexes is also discussed.

A novel peptide fluorescent probe based on different fluorescence responses for detection of mercury species and hydrogen sulfide

Mercury species, including inorganic and organic mercury, are extremely harmful to organisms and the environment. Therefore, it is of great significance to rapidly and selectively detect and distinguish inorganic mercury ions and organic mercury. This study reports peptide sequences that can act as inorganic mercury ions and organic mercury ligands. The peptide fluorescent probe KEEP ((Dansyl-Glu-Glu)2-Lys-NH2) can detect mercury species by its coordination with the side chain of glutamic acid. Hg2+ can be detected by the “turn-off” fluorescence mode, while CH3Hg+ and PhHg+ can be detected by the “turn-on” fluorescence mode. This probe with different fluorescence response to inorganic and organic mercury is very rare, which makes it possible to distinguish between Hg2+, CH3Hg+ and PhHg+. KEEP-Hg2+ can also detect hydrogen sulfide (H2S) through fluorescence reproduction. KEEP can detect these three analytes without interference from other substances due to excellent selectivity. KEEP also has good biocompatibility and low biotoxicity, and can successfully monitor the accumulation of Hg2+ and S2- in living cells. This probe provides a new approach for the detection of inorganic mercury and organic mercury.

Acidic pH Promotes Refolding and Macroscopic Assembly of Amyloid β (16-22) Peptides at the Air-Water Interface.

Assembly by amyloid-beta (Aβ) peptides is vital for various neurodegenerative diseases. The process can be accelerated by hydrophobic interfaces such as the cell membrane interface and the air–water interface. Elucidating the assembly mechanism for Aβ peptides at hydrophobic interface requires knowledge of the microscopic structure of interfacial peptides. Here we combine scanning force microscopy, sum-frequency generation spectroscopy, and metadynamics simulations to probe the structure of the central fragment of Aβ peptides at the air–water interface. We find that the structure of interfacial peptides depends on pH: at neutral pH, the peptides adopt a less folded, bending motif by forming intra-hydrogen bonds; at acidic pH, the peptides refold into extended β-strand fibril conformation, which further promotes their macroscopic assembly. The conformational transition of interfacial peptides is driven by the reduced hydrogen bonds, both with water and within peptides, resulting from the protonation of acidic glutamic acid side chains.

Lipid-Dependent Titration of Glutamic Acid at a Bilayer Membrane Interface.

The ionization properties of protein side chains in lipid-bilayer membranes will differ from the canonical values of side chains exposed to an aqueous solution. While the propensities of positively charged side chains of His, Lys, and Arg to release a proton in lipid membranes have been rather well characterized, the propensity for a negatively charged Glu side chain to receive a proton and achieve the neutral state in a bilayer membrane has been less well characterized. Indeed, the ionization of the glutamic acid side chain has been predicted to depend on its depth of burial in a lipid membrane but has been difficult to verify experimentally. To address the issue, we incorporated an interfacial Glu residue at position 4 of a distinct 23-residue transmembrane helix and used 2H NMR to examine the helix properties as a function of pH. We observe that the helix tilt and azimuthal rotation vary little with pH, but the extent of helix unraveling near residues 3 and 4 changes as the Glu residue E4 titrates. Remarkably, the 2H quadrupolar splitting for the side chain of alanine A3 responds to pH with an apparent pKa of 4.8 in 1,2-dilauroyl-sn-glycero-3-phosphocholine (DLPC) and 6.3 in 1,2-dimyristoyl-sn-glycero-3-phosphatidylcholine (DMPC), but is unchanged up to pH 8.0 in 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) in the presence of residue E4. With bilayers composed of alkali-stable ether-linked lipids, the side chain of A3 responds to pH with an apparent pKa of 11.0 in the ether analogue of DOPC. These results suggest that the depth dependence of Glu ionization in lipid-bilayer membranes may be steeper than previously predicted or envisioned.

Post-translational modifications in collagen type I of bone in a mouse model of aging.

The fracture resistance of cortical bone and matrix hydration are known to decline with advanced aging. However, the underlying mechanisms remain poorly understood, and so we investigated levels of matrix proteins and post-translational modifications (PTM) of collagen I in extracts from the tibia of 6-mo. and 20-mo. old BALB/c mice (female and male analysis done separately). Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis revealed that the levels of collagen I deamidation at specific asparagine (Asn) and glutamine (Gln) residues significantly increased with age. Other non-enzymatic PTMs such as carboxymethylation of lysine (CML) were detected as well, but the relative abundance did not vary with age. No significant age-related differences in the abundance of hydroxylysine glycosylation sites were found, but hydroxylation levels at a few of the numerous lysine and proline hydroxylation sites significantly changed by a small amount with age. We performed molecular modeling and dynamics (MD) simulations for three triple helical fragments representing collagen I regions with prominent age-dependent increases in deamidation as identified by LC-MS/MS of male extracts. These 3 fragments included deamidated Asn and Gln residues as follows: 1) an Asn428 site of the α2(I) chain in which deamidation levels increased from 4.4% at 6-mo. to 8.1% at 20-mo., 2) an Asn983 site of the α2(I) chain with a deamidation increase from 18.3% to 36.8% with age and an Asn1052 site of the α1(I) chain with consistent deamidation levels of ~60% across the age groups, and 3) a Gln410 site of the α1(I) chain that went from no detectable deamidation at 6-mo. to 2.7% at 20-mo. and a neighboring Asn421 site of the same chain with an age-related deamidation increase from 3.6% to 16.3%. The deamidation levels at these sites inversely correlated with an estimate of toughness determined from three-point bending tests of the femur mid-diaphysis. MD revealed that the sidechains become more negatively charged at deamidated sites and that deamidation alters hydrogen bonding with water along the collagen backbone while increasing water interactions with the aspartic and glutamic acid sidechains. Our findings suggest a new mechanism of the age-dependent reduction in the fracture resistance of cortical bone whereby deamidation of Asn and Glu residues redistributes bound water within collagen I triple helix.

Towards understanding the mode of action of the cytotoxic triosmium carbonyl cluster Os3(CO)10(NCCH3)2: Its reactivity with amino acids and oligopeptides

The reactions of the triosmium carbonyl cluster Os3(CO)10(NCCH3)2 (1) with a number of protected amino acids, tri- and pentapeptides were examined. The results show that reaction with the side chains follow the order: -SH > -COOH > -CONH2, and there is a slight preference for the longer side chain of glutamic acid over aspartic acid. The trends observed are retained under physiological conditions and in the peptides.

Lipid-Dependent Titration of Glutamic Acid at a Membrane Interface

The ionization of the glutamic acid side chain has been predicted to depend on its depth of burial in a lipid-bilayer membrane (Biophys J 94, 3393) but has been difficult to verify experimentally (BBA 1859, 484). To address the issue, we incorporated Glu at position 4 in the transmembrane helix of GWALP23 to yield the peptide acetyl-GGAE4W(LA)6LWLAGA-amide. The parent helix exhibits limited dynamic averaging in bilayer membranes and is tilted from the bilayer normal at a well-defined angle that varies systematically with the membrane thickness (J Biol Chem 285, 31723). Residues A3 and A21 are unwound from the core helix. To detect the titration of residue E4, we labeled the side chain of alanine A3 with deuterium and recorded the 2H NMR spectra for oriented samples of the helix in several lipid-bilayer membranes as a function of pH. Remarkably, the 2H quadrupolar splitting for the side chain of A3 responds to pH with an apparent pKa of 4.8 in DLPC and 6.3 in DMPC but is unchanged up to pH 8.0 in DOPC in the presence of residue E4. With bilayers composed of alkali-stable ether-linked lipids, the side chain of A3 responds to pH with an apparent pKa of 11.0 in the ether analogue of DOPC. These results suggest that the depth dependence of Glu ionization in lipid-bilayer membranes may be steeper than previously predicted or envisioned.

Insight into nucleophilic fragmentation mechanisms by glutamic acid side chain in singly protonated glutathione and related peptidyl ions.

Fragmentation mechanisms of the singly protonated glutathione (γ-ECG) and its synthetic analogue peptides (ECG and PPECG) have been investigated by liquid chromatography tandem-mass spectrometry and theoretical calculations. In the mass spectra, similar fragmentation patterns were observed for γ-ECG and ECG, but a completely different one was found in the case of PPECG. The E-C amide bond cleavage is the predominant pathway for the fragmentation of γ-ECG and ECG, whereas the additional N-terminal prolyl residues in PPECG significantly suppress the E-C amide bond cleavage. Theoretical calculations reveal that the fragmentation efficiencies of the E-C bonds in the protonated γ-ECG and ECG are much higher than that in the protonated PPECG, being attributed to their lower barriers of the potential energy; clearly the introduction of two prolyl residues can increase substantially the potential energy barrier. In the proposed mechanism, the protonated E-C amide bonds in the three peptides are first weakened followed by a nucleophilic addition by the glutamyl carboxyl oxygen atom in side chain, leading to the breaking of the E-C amide bonds. However, the processes of E-C bond fragmentation for three protonated analogs were not collaborative. Protonated amide bonds first fragment, then the nucleophilic addition by the side chain of glutamyl carboxyl oxygen atom takes places. On the other hand, the prolyl residues in PPECG can largely diminish the nucleophilic addition, resulting in a much lower efficiency of its E-C amide bond breaking. Distance analysis indicates that breaking the E-C amide bonds in the protonated γ-ECG, ECG, and PPECG ions could not occur without the assistance from the nucleophilic attack, highlighting an asynchronous collaborative process in the bond breakings.

Cloning and Expression of Genes for Biodegrading Nodularin by Sphingopyxis sp. USTB-05.

Biodegradation is efficient for removing cyanobacterial toxins, such as microcystins (MCs) and nodularin (NOD). However, not all the microbial strains with the microcystin-biodegrading enzymes MlrA and MlrC could biodegrade NOD. Studies on genes and enzymes for biodegrading NOD can reveal the function and the biodegradation pathway of NOD. Based on successful cloning and expression of the USTB-05-A and USTB-05-C genes from Sphingopyxis sp. USTB-05, which are responsible for the biodegradation of MCs, the pathway for biodegrading NOD by these two enzymes was investigated in this study. The findings showed that the enzyme USTB-05-A converted cyclic NOD (m/z 825.4516) into its linear type as the first product by hydrolyzing the arginine and Adda peptide bond, and that USTB-05-C cut off the Adda and glutamic acid peptide bond of linearized NOD (m/z 843.4616) and produced dimeric Adda (m/z 663.4377) as the second product. Further, based on the homology modeling of enzyme USTB-05-A, site-directed mutants of USTB-05-A were constructed and seven crucial sites for enzyme USTB-05-A activity were found. A complete enzymatic mechanism for NOD biodegradation by USTB-05-A in the first step was proposed: glutamic acid 172 and histidine 205 activate a water molecule facilitating a nucleophilic attack on the arginine and Adda peptide bond of NOD; tryptophan 176 and tryptophan 201 contact the carboxylate side chain of glutamic acid 172 and accelerate the reaction rates; and histidine 260 and asparagine 264 function as an oxyanion hole to stabilize the transition states.

β-sheet assembly in amyloidogenic glutamic acid nanostructures: Insights from X-ray scattering and infrared nanospectroscopy.

Glutamic acid-rich peptides are crucial to a variety of biological processes, including glutamatergic neurotransmission and immunological defense. Glutamic acid sequences often exhibit unusual organization into β2 -type sheets, where bifurcated H bonds formed between glutamic acid side chains and NH in amide bonds on adjacent β-strands play a paramount role for stabilizing the molecular assembly. Herein, we investigate the self-assembly and supramolecular structure of simplified models consisting of alternating glutamic acid/phenylalanine residues. Small-angle X-ray scattering and atomic force microscopy show that the aggregation pathway is characterized by the formation of small oligomers, followed by coalescence into nanofibrils and nanotapes. Amyloidogenic features are further demonstrated through fiber X-ray diffraction, which reveal molecular packing according to cross-β patterns, where β-strands appear perpendicularly oriented to the long axis of nanofibrils and nanotapes. Nanoscale infrared spectroscopy from individual nanoparticles on dried samples shows a remarkable decrease of β2 -sheet content, accompanied by growth of standard β-sheet fractions, indicating a β2 -to-β1 transition as a consequence of the release of solvent from the interstices of peptide assemblies. Our findings highlight the key role played by water molecules in mediating H-bond formation in β2 -sheets commonly found in amyloidogenic glutamic acid-rich aggregates.

Mass Spectrometry Identifies Isopeptide Cross-Links Promoted by Diethylphosphorylated Lysine in Proteins Treated with Chlorpyrifos Oxon.

Exposure to chlorpyrifos at doses that do not inhibit acetylcholinesterase can be followed by chronic illness in adults and developmental deficits in children. A mechanism to explain these effects is not available. Using mass spectrometry, we have found that chlorpyrifos oxon is a cross-linking agent. Pure proteins incubated with 1.5 mM chlorpyrifos oxon were diethylphosphorylated on lysine and tyrosine. The diethylphospho-lysine reacted with the carboxyl side-chain of aspartic and glutamic acid to form an isopeptide cross-link, releasing diethylphosphate in the process. Of the 14 proteins tested, 9 had cross-links between distinct proteins or between monomers of the same protein, whereas 8 had a cyclic structure created by joining side-chains of nearby residues through an isopeptide bond. The precursor lysine in the isopeptide bond was diethylphosphorylated on the ε-amino group. Tubulin was more susceptible to chlorpyrifos-oxon-induced cross-linking than the other proteins (10 cross-links in tubulin, 2 in human albumin). The role of diethylphospho-tyrosine was not examined. We hypothesize that the protein misfolding and protein cross-linking induced by exposure to chlorpyrifos oxon, via metabolism of chlorpyrifos, could disrupt function, particularly of tubulin, thus leading to chronic illness. Our proposed mechanism is hypothetical until the many questions it raises have been addressed.

Influence of Charged Lipids on Glutamic Acid Containing Transmembrane Helices

Among the crucial components of the cell are transmembrane proteins. These proteins play major roles in many biological activities and are essential for proper cell functioning. An important aspect of these proteins is their respective ionization properties. For glutamic acid, an important consideration is its interaction with lipids in the cell membrane, as little is known about the properties of residues that can titrate from neutral to negative within the hydrophobic environment. In our strictly outlined peptide-lipid system, we substitute a glutamic acid residue in the L14 position of the helical model peptide GWALP23 (acetyl-GGALWLALALALAL14ALALWLAGA-amide). The substitutions of glutamine and aspartic acid will serve as controls. The GWALP23 derivatives are placed in different lipid environments containing negatively or positively charged lipids among the primary zwitterionic lipids of the membrane. The purpose is to create an environment similar to a cell membrane that can also accommodate the glutamic acid side chain. Core alanine resides are labeled with deuterium to determine helix characteristics by solid-state NMR. The peptide-lipid samples will include DMPC doped with 10% charged lipid, as lipid bilayer formation has been confirmed for these lipid mixtures by solid state 31P NMR. The charged lipids will consist of the negatively charged lipid, DMPG, and the positively charged lipid DMTAP. Samples with a 20% charged lipid mixture do not form adequately oriented bilayers with the E14 peptide present. Initially, these experiments will monitor changes in the quadrupolar splittings of the neighboring deuterated alanine residues at positions 15 and 13 as the sample pH is varied. Samples containing E14 and 10% cationic lipid buffered between a pH range of 5.5-8.1 form isotropic peaks in both 31P and 2H-NMR observables. Additional experiments are underway in different lipid mixtures to monitor changes in helix behavior.

Methionine329 in human serum albumin: A novel target for alkylation by sulfur mustard.

Exposure to the vesicant sulfur mustard (SM) may lead to erythema and blistering. Toxicity of SM is hypothesized due to the alkylation of DNA bases and nucleophilic amino acid side chains in proteins (adducts) by forming the hydroxyethylthioethyl (HETE) moiety. Despite its prohibition by the chemical weapons convention, SM still represents a serious threat to military personnel and civilians. Therefore, development and improvement of forensic analytical methods for the verification of SM exposure is of high interest. Protein adducts have been shown to be highly suitable and beneficial biomarkers of poisoning. Herein we present methionine329 in human serum albumin (HSA) as a novel target of SM forming a HETE-methionyl sulfonium ion. The alkylated tetrapeptide LeuGlyMet329 (-HETE)Phe, LGM(-HETE)F, was detected after pepsin-mediated proteolysis and subsequent analysis by microbore liquid chromatography-electrospray ionization-high-resolution tandem-mass spectrometry. Compound identity was confirmed by a synthetic reference. Proteolysis conditions for HSA were optimized towards maximum yield of LGM(-HETE)F and its limit of identification (32.3nM SM in serum) was similar to those of the established HSA-derived biomarkers HETE-CysPro and HETE-CysProPhe (15.6nM SM in serum). Stability of the alkylated Met329 in vitro and in vivo was limited to 5days making this modification a beneficial short-time biomarker. Furthermore, it was found that the HETE-methionyl sulfonium ion can transfer its HETE moiety to the side chain of cysteine and glutamic acid as well as to the N-terminus of peptides and proteins in vitro thus revealing novel insights into the molecular toxicity of SM.

Interaction of Uranium(VI) with α-Amylase and Its Implication for Enzyme Activity.

Because of its chemo- and radiotoxicity, the incorporation of uranium into human body via ingestion potentially poses a serious health risk. When ingested, the gastrointestinal fluids are the primary media to interact with uranium, eventually influencing and even determining its biochemical behavior in the gastrointestinal tract and thereafter. The chemical interactions between uranium and the components of gastrointestinal fluids are, however, poorly understood to date. In this study, the complexation of uranium(VI) (as the uranyl ion, UO22+) with the protein α-amylase, one of the major enzymes in saliva and pancreatic juices, was investigated over a wide range of pH or uranium/α-amylase concentrations covering physiological conditions. Macroscopic sorption experiments suggested a strong and fast complexation of UO22+ to α-amylase between pH 5 and 7. Potentiometric titration was employed to determine the complex stability constants for the relevant UO22+ α-amylase complexes, which is crucial for reliable thermochemical modeling to assess the potential health risk of uranium. Extended X-ray absorption fine structure (EXAFS) spectroscopy revealed that α-amylase is interacting with UO22+ primarily via its carboxylate groups presumably from the aspartic acid and glutamic acid side chains. The effect of UO22+ on the enzyme activity was also investigated to understand the potential implication of uranium for the in vivo functions of the digestive fluids, indicating that the presence of uranium inhibits the enzyme activity. This inhibitory effect can be, however, suppressed by an excess of calcium.

Platinum(II)-glutamic acid dendrimer conjugates: Synthesis, characterization, DFT calculation, conformational analysis and catalytic properties

Abstract A series of platinum(II)- l -glutamic acid dendrimer conjugates having the formula PtCl2(5,5′-Gn(OR)-2,2′-bipyridine); Gn (n = 0, 1, 2), have been synthesized by various amide bond formation methods. The dendrimer complexes were terminated as carboxylic acid (R = H) or methyloxy ester (R = Me). The complexes were characterized by mass spectrometry, elemental analysis, UV-vis, FTIR, 1H-, 13C NMR, 1H-TOCSY, HSQC. The G1(OH) complex aggregate in solution and solid state, giving rise to a rigid conformation for the glutmaic acid side chain. The G2(OH) exists in the “monomeric” form with several conformations for the glutamic acid side chains. Complexes terminated with carboxyl group (R = H) were active catalyst toward hydrogen gas (H2) production by photo-induced splitting of water. The turnover frequency (TOF) for the G1(OH) complex is 14.7 × 1015 s−1, and it exhibits a ∼ 2-fold enhancement in H2 production compared to the unmodified parent complex G0(OH) (TOF = 7×1015 s−1). While G2(OH) complex displayed a lower hydrogen activity (TOF = 3.8 × 1015 s−1) compared to G0(OH). The platinum-glutamic acid dendrimer conjugates showed a correlation between H2 evolution activity and their ability to form aggregates, which driven by the steric hindrance of the peptide residues in the vicinity of Pt-center.

Investigation of Cysteine as an Activator of Side-Chain N→S Acyl Transfer and Tail-to-Side-Chain Cyclization

N→S Acyl transfer is a popular method for the postsynthesis production of peptide C α-thioesters for use in native chemical ligation and for the synthesis of head-to-tail cyclic peptides. Meanwhile thioester formation at the side chain of aspartic or glutamic acids, leading to tail-to-side-chain-cyclized species, is less common. Herein we explore the potential for cysteine to function as a latent thioester when appended to the side chain of glutamic acid. Initial insights gained through study of C-terminal β-alanine as a model for the increased chain length were ultimately applied to peptide macrocyclization. Our results emphasize the increased barrier to acyl transfer at the glutamic acid side chain and indicate how a slow reaction, facilitated by cysteine itself, may be accelerated by fine-tuning of the stereoelectronic environment.

Chemistry at the protein–mineral interface in L-ferritin assists the assembly of a functional (μ3-oxo)Tris[(μ2-peroxo)] triiron(III) cluster

X-ray structures of homopolymeric L-ferritin obtained by freezing protein crystals at increasing exposure times to a ferrous solution showed the progressive formation of a triiron cluster on the inner cage surface of each subunit. After 60 min exposure, a fully assembled (μ3-oxo)Tris[(μ2-peroxo)(μ2-glutamato-κO:κO')](glutamato-κO)(diaquo)triiron(III) anionic cluster appears in human L-ferritin. Glu60, Glu61, and Glu64 provide the anchoring of the cluster to the protein cage. Glu57 shuttles incoming iron ions toward the cluster. We observed a similar metallocluster in horse spleen L-ferritin, indicating that it represents a common feature of mammalian L-ferritins. The structures suggest a mechanism for iron mineral formation at the protein interface. The functional significance of the observed patch of carboxylate side chains and resulting metallocluster for biomineralization emerges from the lower iron oxidation rate measured in the E60AE61AE64A variant of human L-ferritin, leading to the proposal that the observed metallocluster corresponds to the suggested, but yet unobserved, nucleation site of L-ferritin.