Methionine synthesis in Neurospora. The isolation of cystathionine

Among artificially produced biochemical mutants of Neurospora, those which have lost the ability to synthesize methionine form the largest class. At the present writing 87 occurrences of the methionineless character have been observed in this laboratory following treatment of wild type spores with high frequency radiations (1) or mustard gas (2). Methionineless mutants differ from wild type Neurospora in that they fail to grow on a medium containing only sugar, inorganic salts, and biotin, but do grow if, in addition to these constituents, methionine is supplied. In many of the mutants failure of methionine synthesis results from a block in the reduction of sulfate, which, except for a trace of biotin, is the sole source of sulfur in the basal medium. These strains can utilize reduced forms of inorganic sulfur for growth, as well as methionine and other organic sulfur compounds. On the other hand, some of the mutants require organically bound sulfur for growth, an indication that in these strains the block in methionine synthesis comes at a later stage than sulfate reduction. Similar classes of methionine-requiring mutants have been reported in the mold Ophiostoma by Fries (3) and in Escherichia coli by Lampen et al. (4-6)

The d-amino acid oxidase of Neurospora

Among artificially produced mutants of the mold Neurospora have been found strains lacking the ability to synthesize specific amino acids (1, 2). In the course of biochemical and genetic studies of this group of mutants it was observed that some of the mutants, e.g. those deficient in methionine, leucine, and arginine,(1) are able to utilize racemic mixtures of the amino acids with the same efficiency as the l, or physiologically occurring, forms. In the cases of the leucine- and the methionine-requiring mutants it was also possible to show utilization of the ar-keto analogues. It thus appeared possible that the mode of conversion of the d to the l isomers consists in oxidative deammation, followed by resynthesis. A study was therefore undertaken to test the ability of Neurospora to oxidize the “unnatural” optical isomers of the amino acids. It was found that extracts of the mold contain a d-amino acid oxidase similar in its action to the d-amino acid oxidase of mammalian kidney and liver (3). This finding supports the above hypothesis for the conversion of the d- to the l-amino acids. Since it appears that the d-amino acid oxidase has not been previously described in fungi, a number of experiments were performed on the Neurospora enzyme, the results of which are reported here

The biological significance of the search for extraterrestrial life

Biological significance of genetic definition of life in studies of extraterrestrial lif

Biochemical genetics

The field under review is growing so rapidly that it is impossible to cover more than a sampling of recent papers in the allotted space. Important subjects such as the genetics and chemistry of viruses and certain topics in bacterial genetics have had to be omitted, while others have not received the treatment they deserve. Studies of a primarily biochemical nature in which mutants have been employed as tools have been reviewed, as is customary, although it is recognized that their genetic interest lies chiefly in their providing materials for the further study of gene action

The ornithine cycle in Neurospora and its genetic control

It has been emphasized by Haldane (1) that for studies of intermediary metabolism "the new science of genetics furnishes a very powerful method." Such a method is founded upon the general premises that genes control many of the chemical reactions within an organism, and that gene mutations by blocking a reaction chain at various points may, in effect, resolve a metabolic process into some of its constituent stages. For instance, the genetics of such diseases as alcaptonuria and cystinuria have elucidated certain problems in human metabolic processes (2), and studies in the genetics of plant pigments have increased the knowledge of of the biochemistry of anthocyanins (3). But the study of metabolism by way of genetic differences in naturally occurring populations is limited not only by the low rate of mutation but also by the lethal character of most mutations of genes controlling vital functions. By increasing the mutation rate of an organism, through irradiation or otherwise, it is possible to create a number of genetic blocks at various steps in the syntheses of substances or in other processes of metabolism. The problem of preserving mutations ordinarily lethal has been met by Beadle and Tatum (4) in a general course of procedure developed around work with the ascomycetous mold Neurospora. The wild type of this organism is able to carry out all the syntheses essential to its normal growth and reproduction if biotin, inorganic salts, and a suitable source of carbon are available. Strains of Neurospora are irradiated with x-ray or ultraviolet rays on the assumption that mutations will be induced in genes controlling the syntheses of such substances as vitamins and amino acids. Mutant strains of this kind cannot grow on merely inorganic salts, sugar, and biotin, "minimal medium," but can be expected to grow if the product of the blocked synthesis is added to the minimal medium. From irradiated Neurospora there has been isolated in this laboratory a series of mutant strains which require for growth the presence of arginine in the culture medium. A study of the specific biochemical characteristics of members of this group of mutants has made it possible to demonstrate in Neurospora crassa an ornithine cycle similar to that proposed by Krebs and Henseleit (5) as occurring in mammalian liver, and to assign various steps in the cycle to the influence of particular single genes. To our knowledge the ornithine cycle has not previously been demonstrated in plants

Experiments on the carboxylase of pea roots

It is known that vitamin B1 is a growth factor for numerous bacteria and fungi including the yeasts (see the summary in Koser and Saunders (1938)). It has also been demonstrated that vitamin B1 is essential for the growth of the isolated roots of higher plants (Bonner, 1937; Robbins and Bartley, 1937). Because of this general vitamin B1 requirement of living organisms, it would seem a priori probable that the vitamin plays a role in some basic cellular process. That this is indeed the case was shown conclusively by the work of Peters and coworkers (see Peters and O’Brien (1938)) and of Lohmann and Schuster (1937). The latter workers found that the prosthetic group of yeast carboxylase is vitamin B1 pyrophosphate. In the case of yeast, vitamin B1 is, then, a constituent of a respiratory enzyme and vitamin B1 pyrophosphate is hence commonly referred to as “cocarboxylase,” a terminology used throughout this paper. Although considerable information is available concerning the rôle of vitamin B1 as a growth factor for roots, there is little known about the carboxylase of such roots. The present work was undertaken with the hope of elucidating possible relationships between vitamin B1 and the carboxylase of pea roots

Growth inhibition of neurospora by canavanine, and its reversal

Canavanine, an amino acid from jack beans, was discovered by Kitagawa and coworkers in 1929 (1, 2). The substance is not combined in the proteins of the seed, but occurs in the free state, and makes up 2.5 per cent of the dry weight of jack beans (3). In a series of papers available to the authors for the most part in abstract only, the Japanese workers have reported extensive investigations into the chemistry and physiology of the substance. The structure of canavanine was established by Gulland and Morris (4) and by Kitagawa and Takani (5) as NH2•C(:NII)•NII•O•CH2•CH2•CHNH2•COOH. Natural canavanine is of the L configuration (6)

Physiological Aspects of Genetics

A considerable amount of evidence indicates that desoxyribonucleic acid is capable of duplicating itself, a property also possessed by genes. (By a self-duplicating material, we mean one which plays some essential role in its own production.) Watson & Crick (1) have proposed a new structure for desoxyribonucleic acid which not only takes into account the existing analytical and x-ray diffraction data but also seems capable of explaining the mechanism of duplication. Their model consists of two helical chains coiled around the same axis, the purine and pyrimidine bases on the inside, the phosphate groups on the outside. The chains are held together by hydrogen bonds between the bases, the adenine residues of either chain being bonded specifically to thymine in the other, and similarly guanine to cytosine. The sequence of bases along one chain is not restricted, but once fixed the sequence along the other chain is determined. This complementarity, which is the most novel feature of the structure, suggests that duplication takes place by separation of the two chains, followed by the synthesis of its complement alongside each chain. The model is supported by recent x-ray diffraction studies (2, 3)

The L-amino acid oxidase of Neurospora

In 1944 one of us described a D-amino acid oxidase in extracts of Neurospora crassa (1). Except for slight activity against L-glutamate, no oxidation of L-amino acids was observed. Recently a means for inducing the formation of a soluble L-amino acid oxidase by the mold was reported by Bender, Krebs, and Horowitz (2). This is accomplished by reducing the biotin content of the basal medium (3) from the 5 γ per liter usually employed to 0.25 γ per liter. When cultured in such a medium, Neurospora produces an active L-amino acid oxidase which can be demonstrated both in extracts and in the medium. Reduction of the biotin level produces no effect on the D-amino acid oxidase, which is still found in extracts but not in the culture medium. The activity of the L-oxidase toward thirty-eight amino acids has been investigated by Bender and Krebs (4). The initial purpose of the present investigation was to explore certain possibilities for a genetic study of the enzyme. Of special interest was the fact, cited by Bender et al. (a), that the oxidase could not be detected in all of the wild type strains tested. The question arose as to whether this is due to the genetic inability of certain strains to form the enzyme. In connection with the investigation of this problem a general survey of the properties of the enzyme was carried out, together with a preliminary study of the mechanism of the biotin effect and of the adaptive formation of the enzyme. The results of these studies are reported below. Simultaneously with our investigation and independently of it, Dr. K. Burton (5), working in Professor Krebs’ laboratory, has carried out a study of the Neurospora L-amino acid oxidase. Where our respective studies overlap mutual confirmation was obtained in most essential points. We wish to thank Dr. Burton for permission to read his manuscript before publication

The Origin of Life

The origin of life is in a sense a genetic problem, for, as H. J. Muller pointed out many years ago, the essential attribute that identifies living matter is its capacity to replicate itself and its variants (1). Because this uniquely biological property has its physical basis in proteins and nucleic acids, the goal of modern work on the origin of life is to discover the manner of origin of these polymers and of the interactions between them that constitute the genetic mechanism. In attempting to review this subject in a limited space, we cannot undertake an exhaustive treatment. Rather, we summarize work published principally since 1970 in the following areas, with emphasis on those aspects that are of greatest current interest: 1. precambrian paleontology, 2. chemical evolution of genetically important monomers, 3. prebiotic dehydration-condensation reactions, 4. organic compounds in meteorites and interstellar space, and 5. biological exploration of the planets. A large number of review articles (2-5), critical and theoretical discussions (6-8), books (9-16), and conference proceedings (17-21) dealing with the origin of have appeared in recent years. In addition, a new serial, the Journal of Molecular Evolution, publishing papers on this and related subjects, appeared in 1971; the journal Space Life Sciences has been renamed "Origins of Life," and a society, the International Society for the Study of the Origin of Life, was recently founded