Spectral properties of conformational motion in proteins

The conformational dynamics of large fragments of protein structure within the framework of a generalized model of bounded diffusion (GMBD) have been considered. This model is a development of the known ordinary model of bounded diffusion (MBD). The latter model assumes that the driving force of motion, which is at the same time a source of friction, has negligibly small correlation time (white noise). In contrast to the MBD, the GMBD takes into account the finite character of friction correlation time, i.e. memory friction effects. Two different mechanisms of friction for the fragment are considered: (1) friction by rapid density fluctuations due to the harmonic vibrational motion of atoms and (2) friction by slow encounters with surrounding fragments of protein structure. The present theory shows that at high frequencies the power decay of the spectrum which takes place in the MBD is replaced by an exponential one. In the narrow intermediate frequency range the results of the GMBD coincide with those of the MBD. At low frequencies the results of the two models differ quantitatively, though their qualitative behaviour is similar.     

Reaction of nitric oxide with heme proteins: studies on metmyoglobin, opossum methemoglobin, and microperoxidase

Kinetic and EPR studies show that the first step in the reaction of NO with ferric myoglobin, opossum hemoglobin, and microperoxidase is the reversible formation of the H-NO complex: H + NO in equilibrium H-NO (where H = Mb+, or Hb+ OP, or MP+). The NO-combination rates are markedly affected by the presence or absence of the distal histidine. The distal histidine significantly reduces the NO-combination rates, perhaps by interaction between the distal histidine and the ferric iron. Thus the beta-chains of Hb+ OP and metmyoglobin show similar combination rates. In the absence of a distal histidine, the NO-combination rates in the alpha-chains of Hb+ OP are much faster and similar to those observed for the five-coordinate heme in microperoxidase. The loss of a water molecule from the six-coordination site is assumed to be the rate-limiting step.     

Comparative aspects of methemoglobin formation and reduction in opossum (Didelphis virginiana) and human erythrocytes

Glucose-depleted, nitrite-treated opossum erythrocytes effectively reduce methemoglobin in an environment of physiological saline and added glucose does not accelerate the rate of reduction. In autologous plasma or 25 mM phosphate-buffered saline pH 7.4, added glucose significantly accelerates methemoglobin reduction in glucose-depleted, nitrite-treated opossum erythrocytes. Human red cells require added glucose to carry out reduction of methemoglobin and increased phosphate concentration or autologous plasma does not alter the rate of this process. Within the opossum red cell in vitro, autooxidation of hemoglobin proceeds at a much slower rate than that observed in human erythrocytes.     

Nitric oxide induced conformational changes in opossum hemoglobin.

Opossum hemoglobin assumes a T quaternary structure upon NO ligation in the absence of organic phophates at pH 6.7. In addition, stripped opossum hemoglobin exhibits a low oxygen affinity when compared to human hemoglobin and a pH-dependent heme-heme interaction with an n value of 2.14 at pH 7.0 and 2.46 at pH 7.35. These observations indicate that opossum hemoglobin may have a destabilized oxy structure when compared to hemoglobin A due to differences in primary structure. Thus, the strong trans ligand effect of nitric oxide is able to disrupt the proximal histidine-iron bond in the alpha-hemes triggering a conformational transition to the T state. Absence of a distal histidine in the alpha-subunits and, therefore an impaired donor acceptor interaction with the sixth ligand, could contribute to the lack of stability of the R quaternary structure in opossum nitrosylhemoglobin. The reduced oxygen affinity of opossum hemoglobin may be compensated for by other physiological factors such as a reduced phosphate effect.     

The spatial distribution of physical, chemical, energetic and conformational properties of amino acid residues in globular proteins

An attempt is made to obtain information as to the spatial distribution of amino acid residues in globular proteins in terms of their chemical, physical, energetic and conformational properties. The crystallographic data on twenty-one protein molecules form the basis for the study. The properties of the residues, namely, hydrophobicity, polarity, acidity, molecular weight, bulkness, chromatographic index, refractivity, short/medium/long-range energetics, and powers to adopt a-helical, extended and bend structures are analysed by dividing the protein globule into six concentric shells containing equal numbers of residues. The results show that the decisive factor in determining the spatial position of a residue in a protein molecule is of composite nature involving a compromise between the various properties of the residue. The observed deviations from the general hydrophobic interior and hydrophilic exterior in globular proteins are nicely brought out. A valuable clue is obtained as to the directional properties of α-helical and extended structure segments in proteins. The relative buried, exposed and intermediate characters of the residues in protein globules are obtained in a very realistic approach.     

Studies on the energy metabolism of opossum Didelphis virginiana erythrocytes--III. Metabolic depletion with 2-deoxyglucose markedly accelerates methemoglobin reduction in opossum but not in human erythrocytes

1. Glucose-depleted, nitrite-treated erythrocytes reduce ferriheme in vitro in an environment 100 mM to 2-deoxy-D-glucose at a rate of 2.4 microM/ml cells/hr (opossum) and 0.37 microM/ml cells/hr (human). 2. During the process of methemoglobin reduction the breakdown of adenine ribonucleotides is more rapid in opossum (0.9 microM/g hg/hr) than in human (0.36 microM/g hg/hr) erythrocytes. 3. Radiolabelled ribose from [U-14C] ATP is catabolized exclusively to [14C] lactate in opossum, and to [14C] pyruvate and [14C] lactate in human red cells.     

The extensor assembly in two species of opossum,Philander opossum and Didelphis marsupialis

In considering primate and hominoid phylogeny, the fundamental position assigned to opossums is explained partially by the characteristic morphology of their hands and feet. One of the main functional features of the human hand is the ability to make a stabilized arch of the finger. Because the extensor assembly plays a key role in establishing an arched finger, the extensor systems of the digits of both the hands and feet were studied in two species of opossum, Philander opossum and Didelphis marsupialis. In the foot, two extensor tendons join in each toe to form one tendinous plate, which inserts onto the base of the second phalanx. Lumbricals join this plate along the tibial side, and interosseus insertions are found, although a true interosseus wing is lacking. At the proximal interphalangeal level, a terminal tendon takes its origin from this tendinous plate. This terminal tendon is oval in cross-section and contains elastic structures. Oblique bands arise from this terminal tendon and run proximally along the proximal interphalangeal joint inserting onto the base of the first phalanx. There are elastic structures in the flexor tendon on the dorsal side near its site of insertion. In the hand, the main extensor tendons are arranged differently and the interossei contribute substantially to the extensor assembly. Otherwise, the extensor assembly of the hands and feet are quite similar. The function of the so-called paratendinous intravaginal flexors is discussed as are evolutionary aspects of the extensor assembly.     

Circular dichroism and conformational properties of modeccin,a toxic protein

According to its circular dichroism (CD) spectrum, modeccin, a toxic lectin from the roots of the South African plantModecca digitata, is structurally similar to the ricins and abrins. In nearly neutral and weakly alkaline solutions (pH 7.6–9.0) the CD spectra of modeccin displayed a positive CD band at 190–195 nm and a negative band at 210–220 nm, indicating the presence of some α-helix and β-sheet structures. In the near-ultraviolet zone, we observed positive CD bands at 232 and 245 nm and weak negative bands at 285 and 293 nm. In more strongly alkaline solutions of pH 9.5–10.2 the CD bands in the farultraviolet zone were not affected, but the CD band at 232 nm diminished and the CD band at 245 nm was enhanced. These transitions were reversible. At pH 11.2–11.5 the CD band at 232 nm disappeared completely, and the CD bands in the far-ultraviolet diminished. The CD bands at 285 and 293 nm were affected very little by the alkali, and these bands were assigned to buried tryptophan side chains. Sodium dodecyl sulfate and 2,2,2-trifluoroethanol disorganized the tertiary structure of modeccin and reconstructed the secondary structure into a new form with a higher helix content than in the native protein.     

Structural fluctuations and conformational entropy in proteins: entropy balance in an intramolecular reaction in methemoglobin.

The reversible intramolecular binding of the distal histidine side chain to the heme iron in methemoglobin is of special interest due to the very large negative reaction entropy which overcompensates the large reaction enthalpy. It may be considered as a prominent example of the ability of proteins (including enzymes) to provide global entropy in a local process. In this work new experiments and model calculations are reported which aim at finding the structural elements contributing to the reaction entropy. Geometrical studies prove the implication of the 20 residue E-helix being shifted by more than 2 A. Vibrational entropies are calculated by a procedure derived from the method of Karplus and Kushik. It turns out that neither the histidine alone nor the complete E-helix contribute more than 15 per cent of the required entropy.     

Spectral properties of perselenosulfide

The riductive scission of a diselenide (selenocystamine) produced by disulfide gives the perselenosulfide, a new compound. Its formation kinetics, carried out a several pH values, were recorded spectrophotometrically at 375 nm. The lability of perselenosulfide bond by cianolysis has been demonstrated.     

Functional and conformational properties of mouse hemoglobins

Hemoglobins from four strains of mice (C3H/SW, DBA/2J, C57BL6/Kh and A.TH) examined showed pH-dependent heme-heme interactions. The oxygen affinity and cooperativity are reduced at acidic pH. The oxygen equilibrium parameters increase as a function of increasing pH and at physiological pH values they are similar to the corresponding values of human hemoglobin A. The nitrosyl derivatives of these mouse hemoglobins undergo a quaternary structural transition to the T state in going from pH 7.0 to 6.0. These functional and conformational properties are indicative of destabilised oxy structures of mouse hemoglobins at acidic pH. This study also confirms that the cysteine residue at beta 13(A10) position has no influence on the oxygen equilibrium properties or conformation of the molecule.