The chemistry of vitamin B12. The coordination of biologically important molecules

The following equilibrium constants (given as logK in units of m⁻¹) were determined for the substitution of co-ordinated H₂O in aquocobalamin by glycine (bound through N) 5.8, cysteine (bound through S) 6.0 or 8.3, depending on the value chosen for the pK of the thiol group, and phenolate 2.9. The spectrum of the phenolate cobalamin shows an additional intense absorption band at 468nm with a molar extinction coefficient of 1.1×10⁴, which is assigned to a charge transfer from the phenolate to the cobalt ion. Equilibrium constants have also been determined for the equilibria between adenylcobamide cyanide and CN⁻, HO⁻ and H⁺, which show that the adenine is more easily displaced by CN⁻ and HO⁻ than is 5,6-dimethylbenziminazole in vitamin B₁₂, but can be protonated by acid while still remaining co-ordinated to the cobalt. It is shown that in the binding of corrinoids to proteins and polypeptides the formation of hydrogen bonds is far more important than co-ordination by the metal.     

Phenylketonuria—Experience at One Center in the First Year of Screening in California

One year''s experience with phenylketonuria during the calendar year 1966, the first year for compulsory newborn screening in California, was reviewed. The over-all prevalence rate from reported cases in California during this period was one case per 19,500 persons tested. Fifty-seven persons suspected of having pku were evaluated, and 25 of them were determined to be phenylketonuric. Eleven of the 25 were infants in whom the abnormality was detected through the newborn screening program or because it was detected in a sibling through a screening program. All the newborn phenylketonuric patients were developing normally at the time of last report (although the follow-up periods were short).In nine of the other children, pku was detected because they were retarded. Five retarded children who were diagnosed as phenylketonuric at another clinic were given dietary assistance.Five additional infants had elevated serum phenylalanines but did not have the classic biochemical findings of pku and are being evaluated further. Nine infants with positive screening tests exhibited biochemical and clinical findings consistent with transient tyrosinemia. Eighteen other children were evaluated and found to have no metabolic abnormality.The newborn screening program for pku is of decided benefit in early identification of a group of infants who have a high rate of potentially serious metabolic disease. Early identification permits treatment soon enough to prevent mental retardation. Newly identified patients should be evaluated in a medical setting capable of careful pediatric, biochemical and nutritional surveillance.     

The purine and pyrimidine metabolism of Tetrahymena pyriformis

The metabolism of purines and pyrimidines by the ciliated protozoan Tetrahymena was investigated with the use of enzymatic assays and radioactive tracers. A survey of enzymes involved in purine metabolism revealed that the activities of inosine and guanosine phosphorylase (purine nucleoside: orthophosphate ribosyltransferase, E.C. 2.4.2.1) were high, but adenosine phosphorylase activity could not be demonstrated. The apparent K_m for guanosine in the system catalyzing its phosphorolysis was 4.1 ± 0.6 × 10^?3 M. Pyrophosphorylase activities for IMP and GMP (GMP: pyrophosphate phosphoribosyltransferase, E.C. 2.4.2.8), AMP (AMP: pyrophosphate phosphoribosyltransferase, E.C. 2.4.2.7), and 6-mercaptopurine ribonucleotide were also found in this organism; but a number of purine and pyrimidine analogs did not function as substrates for these enzymes. The metabolism of labeled guanine and hypoxanthine by intact cells was consistent with the presence of the phosphorylases and pyrophosphorylases of purine metabolism found by enzymatic studies. Assays for adenosine kinase (ATP: adenosine 5'-phosphotransferase, E.C. 2.7.1.20) inosine kinase, guanosine kinase, xanthine oxidase (xanthine: O₂ oxidoreductase, E.C. 1.2.3.2), and GMP reductase (reduced-NADP: GMP oxidoreductase [deaminating], E.C. 1.6.6.8) were all negative. In pyrimidine metabolism, cytidine-deoxycytidine deaminase (cytidine aminohydrolase, E.C. 3.5.4.5), thymidine phosphorylase (thymidine: orthophosphate ribosyltransferase, E.C. 2.4.2.4), and uridine-deoxyuridine phosphorylase (uridine: orthophosphate ribosyltransferase, E.C. 2.4.2.3) were active; but cytidine kinase, uridine kinase (ATP: uridine 5'-phosphotransferase, E.C. 2.7.1.48), and CMP pyrophosphorylase could not be demonstrated.     

Genetic Basis of Colicin E Susceptibility in Escherichia coli I. Isolation and Properties of Refractory Mutants and the Preliminary Mapping of Their Mutations

Colicin E-resistant mutants were isolated in Escherichia coli K-12 which, although still apparently possessing the E receptor and adsorbing colicin, were nevertheless insensitive (refractory) to its effect. Eight phenotypic groups were obtained, but some mutants from three of these groups were all shown to map at gal, whereas a second refractory locus, giving resistance to E1 alone, mapped close to thy. It is suggested that the successful fixation of any of the three distinct colicins of group E may involve a dual role for the cell surface "receptor," the first for the binding of the protein and the second for the correct orientation of the bound molecule relative to the cytoplasmic membrane. The majority of the refractory mutants isolated may derive from changes in components concerned with the second of these receptor functions. Two groups of mutants, however, refractory to only E1 or E2, probably reflect changes in the intracellular transmission systems which specifically mediate the effects of these two colicins, the changes not allowing transmission through the cytoplasmic membrane to the respective targets of the colicins. The E1 adsorption site was shown to be distinct from that for E2 and E3, indicating an early separation of the colicin E transmission systems.