P2 Molecule Research Articles

Molecular analysis of the P2 porin protein of nontypeable Haemophilus influenzae

The P2 porin protein is the most abundant outer membrane protein (OMP) of nontypeable Haemophilus influenzae (NTHI) and shows extensive antigenic heterogeneity among strains. To study the molecular basis of this heterogeneity, the DNA sequences of the genes encoding the P2 proteins of three unrelated strains of NTHI were determined, and restriction fragment length polymorphisms around the P2 genes of 35 strains were analyzed. The deduced amino acid sequences of the P2 genes from the three strains of NTHI revealed four major (12 to 35 amino acids long) and several smaller (2 to 7 amino acids) hypervariable regions in each protein. The major variations occurred in identical portions of the genes, and these regions showed a high antigenic index and surface exposure probability in computer modeling analysis. Differences in the molecular mass of the P2 protein correlate with differences in the size of the variable region in each strain. Oligonucleotide primers suitable for amplification of the P2 genes by polymerase chain reaction were developed. Restriction fragment length polymorphism analysis showed marked heterogeneity in and around the ompP2 locus of 35 NTHI strains. These results contrast with the high degree of conservation of the P2 genes in H. influenzae type b strains. We conclude that the molecular mass and antigenic heterogeneity of the P2 molecule of NTHI is due to variations in gene sequence that are clustered primarily in four large hypervariable regions of the gene.

Accurate STF HF wave functions from B to Ca and accurate STF HF wave functions for N2 and P2

Accurate Slater type function (STF) Hartree–Fock (HF) wave functions are calculated from B to Ca. The STF’s have a form of rne−γr and power (n) of r is carefully chosen. The total atomic energies agree with those of numerical HF (NHF) within the errors of 3×10−6 and 1×10−5 a.u. for B to Ne and for Na to Ca, respectively. It will be also shown that for the N2 molecule, the present atomic STF HF set augmented with four 3d, three 4f, and one 5g STF polarization functions gives the total energy of −108.9937 a.u. at R=2.068 a.u. which is quite close to that of NHF, −108.9939 a.u. It is found that one 5g type polarization function is indispensable for getting the accurate HF value. The near HF wave function for the P2 molecule, which has the same quality as that of N2, will be also shown in the present work. The basis sets for N2 are applied to configuration interaction calculations in order to see the role of the polarization functions on the correlation effects.

The complete amino acid sequence of the rabbit P2 protein.

P2 protein is a small basic protein (Mr = 14,820) found in peripheral nerve myelin and spinal cord myelin. There is now overwhelming evidence that P2 protein is the crucial antigen involved in the induction of experimental allergic neuritis, an autoimmune disease of the peripheral nervous system. The complete amino acid sequence of rabbit P2 protein was derived by sequence analysis of cyanogen bromide peptides and peptides obtained by proteolysis using Staphylococcus aureus V8 enzyme, trypsin, or clostripain. There are 131 amino acids and an excess of the basic amino acids lysine and arginine; histidine is absent. There are 3 highly hydrophobic regions in the P2 molecule. Probability analysis of the sequence predicts a high degree of beta structure, essentially in agreement with CD data.

Allergic neuritis: Phospholipid requirement for the disease-inducing conformation of the p2 protein

The P2 protein, a small, highly ordered basic protein of peripheral nerve myelin, is a potent inducer of allergic neuritis in rats when complexed with phospholipids such as phosphatidylserine. lsolated P2 protein administered without lipid is a poor neuritogen, and if first oxidized with performic acid, aminoethylated in 8 M urea or heat denatured, it loses nearly all activity. When the aminoethylated or oxidized forms are combined with phosphatidylserine, however, they recover essentially full neuritogenic activity. Complexing with lipid also greatly enhances the activity of the heat denatured form. Spleen cells sensitized to the aminoethylated and heated forms of P2 protein show a pronounced mitogenic response to either of these forms as well as to the P2 protein itself, but only when sensitization is initiated with the lipid complex. These data indicate that the lipid complex reverses the distortion acquired by chemical treatment or denaturation and converts the P2 molecule into a conformation approximating that of the native P2 protein in myelin. These studies imply that the neuritogenic domain, while highly sensitive to denaturing conditions, requires interaction with phospholipids in order to attain the most favourable conformation for inducing a cell-mediated response that leads to disease.

Radioimmunoassay for the P2 protein of bovine peripheral nerve myelin.

The P2 protein of bovine nerve root myelin was radiolabeled with 125I in homogeneous solution by the chloramine-T method and purified by gel filtration on Sephadex G-75 to obtain monomeric 125I-P2. Antigen-antibody complexes were isolated by the silica gel, double antibody, or polyethylene glycol techniques by using rabbit antibody to P2. As little as 1 ng/ml P2 could be detected. The RIA was used to measure the P2 content in nerve tissue and isolated myelin. The presence of P2 in spinal cord as well as in the peripheral nervous system was confirmed. Peptides isolated by CNBr digestion of the P2 protein were tested in the RIA. CN-1, comprising 80 to 90 residues from the interior of the molecule displayed complete immunologic cross-reactivity with intact P2. Neither CN-2, representing 18 amino acids from the COOH terminal, nor CN-3, representing 20 amino acids from the NH2 terminal, showed cross-reactivity. Since the major determinant for experimental allergic neuritis in the rabbit is located in peptide CN-2, our present data suggest that the major neuritogen and the major determinant(s) for humoral antibody response in this species may be at different locations within the P2 molecule.

HeI photoelectron spectrum of the P2(X 1? + g ) molecule

The HeI photoelectron spectrum of the P2(X1∑+g) molecule has been recorded and interpreted with the aid of ab initio and semi-empirical molecular orbital calculations. Three ionic states of the diphosphorus molecule are found to be accessible with HeI radiation. Comparison is made between the properties of these states and the corresponding well established ionic states of nitrogen.

Spectre d'émission du radical P2 : étude de la transition b′3Σu−–X1Σg+

The emission spectrum of the b′3Σu−–X1Σg+ system between 3540–4375 Å and ascribed previously by Mrozowski and Santaram to the a3Σu+–X1Σg+ transition of the P2 molecule, has been photographed under high resolution. The rotational analysis of 7 bands has been carried out and allowed us to determine the rotational constants of the vibrational levels ν′ = 0, 1, and 2 as well as the spin splitting constants λ′ and γ′. The nature of the upper state has been identified as a 3Σu− state, the electronic transition being analogous to the Ogawa–Tanaka–Wilkinson system of N2.

Estimated potential energy curves of diatomic molecules: a simplified procedure

A simplified method for evaluating f and g involved in the calculation of rmax and rmin for the electronic states of diatomic molecules, whose energy terms cannot be represented by a quadratic in v+1/2, is discussed. The method consists of dividing the observed vibrational term system into segments, each segment consisting of two vibrational levels, which can be represented by a quadratic in v+1/2 by the proper choice of constants omega i and omega xi, and thereby using the expressions of f and g derived by Rees for the quadratic equation, but with a slight modification. As a check, the potential energy curve of the B 3 Pi 0u+ state of the 79Br81Br molecule is constructed and compared with that calculated by Rees' elliptic function and RKRV methods. Further, the classical turning points are calculated for a few vibrational levels of the X 2 Sigma g+ state of N2+ and compared with those obtained by the RKRV method. The agreement is found to be excellent in both cases. Using the new procedure, true potential energy curves are also constructed for the X 1 Sigma g+ and C 1 Sigma u+ states of the P2 molecule.

Rotational analysis of some bands of C1 Σ u + -X1 Σ g + system of P2 molecule involving lowν′,ν″ values

Ten ultraviolet bands of the C1 Σ u + -X1 Σ g + system of P2 involving lowv′ andv″ values have been photographed at dispersion of 0·38 and 0·56 A/mm. and analysed for their rotational structure. While four of these bands were analysed earlier, six of them,viz., 0–10, 1–12, 2–7, 2–14, 4–8 and 6–9 have been analysed for the first time during the present studies. The rotational constants, B v S with lowv″ quantum numbers are obtained from which value of B θ ″ has been derived. The value of B θ ″ is found to be in agreement with the value obtained by Douglas and Rao from their study of A1 Π g-X1 Σ g + bands of P2.

Electronic Spectra of P_2, As_2, and Sb_2 Molecules

The band spectra of diatomic molecules P2, AS2, Sb2 and Bi2 were investigated in the spectral range 2100–10 000 A with a medium-size spectrograph, using a discharge tube developed for studies of forbidden lines. A study of the excitation conditions of various bands shows, among other things, that the bands previously ascribed to As2+ actually are emitted from a low-lying electronic state of the As2 molecule. Several new band systems were found, for some of which a vibrational analysis was carried out. This permits the identification of three new electronic states for the P2 molecule and two new states for the As2 molecule. Four of these states are located below the previously known excited states of these molecules and belong to the triplet term system. A comparison with the N2 spectrum shows that for this group of diatomic molecules the 3Πg state, located above the 3∑u+ state for the N2 molecule, becomes gradually depressed below the 3∑u+ state as the atomic number of the element increases.

ON THE SPECTRUM OF THE P2 MOLECULE

Five bands of the ultra-violet system of P2, with low v′ and v″ values, have been measured under large dispersion and analysed. They serve to determine much more accurately than heretofore possible the rotational constants Be″ and αe″, the moment of inertia Ie″, and the internuclear distance re″ of the P2 molecule in the ground state. The following values have been found: Be″ = 0.3031 cm.−1, αe″ = 0.00138 cm.−1, Ie″ = 92.36∙10−40gm.-cm.2, re″ = 1.895 10−8 cm.