Production of Erythritol by n-Alkane-grown Yeasts

In the course of study on citric acid fermentation by Candida zeylanoides, in which n-alkane (a mixture of C–12 to C–15) was used as the sole source of carbon, we found that a polyol-like substance was accumulated when the medium-pH fell down to below 4.0. This was isolated in crystalline forms and identified as meso-erythritol. Comparing erythritol production among fifty yeast strains, Candida zeylanoides, particularly its glycerol-requiring mutant KY 6166, was found to be an excellent producer.

Erythritol production was also observed with ethanol or acetic acid as the sole carbon source but not with glucose. An efficient condition for large production of erythritol was to keep the medium-pH at low level (2.5 to 4.0) and the concentration of NaCl or KCl at high level (1 to 3%). Under conditions established in this work, more than 55 mg/ml of erythritol was successfully produced in 120 hr incubation in 300-ml flasks, which corresponded to 55% of the alkane used.     

Taq I-generated HLA-DQ αpolymorphism in Japanese patients with narcolepsy

Taq I-generated HLA-DQrestriction fragment length polymorphism was examined in Japanese patients with narcolepsy. All patients were DR2 positive and shared a 6.0 kb fragment, although this fragment was found only in 54 % of the healthy DR2-positive Japanese. This finding added the DQ gene to the list of candidates for the possible narcolepsy-susceptibility gene. In contrast, there was no complete association between narcolepsy and DXrestriction fragment length polymorphism. These findings suggest that a narcolepsy-susceptibility gene is located closer to the DQ locus than to the DX locus.     

Molecular analysis of HLA-B39 subtypes

Serological studies have suggested the presence of a new HLA-B39 subtype (B39.2) in the Japanese population. To identify the new HLA-B39 subtype and compare it with an other HLA-B39 subtype (B39.1), the genes encoding HLA-B39.1 (B ^* 39013) and B39.2 (B ^* 3902) have been cloned from Japanese. We have sequenced these genes and completed the sequence of HLA-B39.1 (B ^*39011 ) gene from a Caucasian that was partially sequenced. Comparison of the sequence data revealed that B ^* 3902 and B ^* 39013 differ by three nucleotide substitutions which result in a two amino acids change at residues 63 and 67, while one silent substitution at codon 312 is found between B ^* 39011 and B ^* 39013. These results suggest that B ^* 3902 has evolved from B ^* 39013 rather than B ^* 39011.The nucleotide sequence data reported in this paper have been submitted to the GenBank nucleotide sequence database and have been assigned the accession numbers M94051 (HLA-B^*39013), M94052 (HLA-B^*39011), and M94053 (HLA-B^*3902).     

Further Evidence for Multiple Forms of an N-Methyl-d-Aspartate Recognition Domain in Rat Brain Using Membrane Binding Techniques

Abstract— Pretreatment with sulfhydryl-reactive agents, such as N-ethylmaleimide and p-chloromercuriphenylsul-fonic acid, invariably resulted in marked inhibition of the binding of dl -(E)-2-amino-4-[³H]propyl-5-phosphono-3-pentenoic acid ([³H]CGP 39653), a competitive antagonist at an N-methyl-d -aspartate (NMDA)-sensitive subclass of central excitatory amino acid receptors, in brain synaptic membranes extensively washed and treated with Triton X-100, but did not significantly affect the binding of L-[³H]-glutamic acid ([³H]Glu), an endogenous agonist. The pre-treatment was effective in reducing the binding of [³H]-CGP 39653 at equilibrium, without altering the initial association rate, and decreased the affinity for the ligand. Pretreatment with sulfhydryl-reactive agents also enhanced the potencies of NMDA agonists to displace [³H]-CGP 39653 binding and attenuated those of NMDA antagonists, but had little effect on the potencies of the agonists and antagonists to displace [³H]Glu binding. The binding of both [³H]CGP 39653 and [³H]Glu was similarly sensitive to pretreatment with four different proteases in Tritontreated membranes, whereas pretreatment with phospho-lipase A₂ or C markedly inhibited [³H]CGP 39653 binding without altering [³H]Glu binding. Moreover, both phospho-lipases not only induced enhancement of the abilities of NMDA agonists to displace the binding of [³H]CGP 39653 and [³H]Glu, but also caused diminution of those of NMDA antagonists. These results suggest that both sulfhydryl-reactive agents and phospholipases may predominantly interfere with radiolabeling of the NMDA recognition domain in a state favorable to an antagonist by [³H]CGP 39653, with concomitant facilitation of that in an agonist-preferring form by [³H]Glu. The possible presence of multiple forms of the NMDA recognition domain is further supported by these data.     

Evolution of glycoproteins as judged by the frequency of occurrence of the tripeptides Asn-X-Ser and Asn-X-Thr in proteins

Summary A statistical analysis of the data tabulated in the Atlas of Protein Sequence and Structure 1972 indicates that the observed frequency of occurrence of the tripeptides Asn-X-Ser and Asn-X-Thr is approximately one third of the expected in eukaryotic proteins, but in prokaryotic proteins the observation agrees closely with expectation. Thus the lowered frequency of these tripeptides found by Hunt and Dayhoff is restricted to eukaryotic proteins. Of all the Asn-X-Ser/Thr sequences examined, those which contain covalently attached carbohydrates are found only in the extracellular proteins of eukaryote. These observations are discussed in relation to the evolution of glycoproteins which seems to have occurred in the ancestor of eukaryotes after the divergence from prokaryotes.     

Acid phosphatase in plant tissues I. Changes in activity and multiple forms in tea leaves and tomato fruit during maturation and senescence

Multiple forms of acid phosphatase varied in tea leaves (Camelliasinensis L.) depending on variety and the age of the leaves.Polyacrylamide gel electrophoresis indicated the presence ofsix multiple forms in extracts of young leaves and seven inextracts of mature leaves of the variety Hatzumomizi. VarietyBenihomare showed five bands in zymograms of young-leaf extractsand six in zymograms of mature-leaf extracts. In addition tothe appearance of the new band, the relative intensity of stainingin two other bands changed during maturation. One decreasedand one increased in intensity. The components were partiallyseparated by CM-cellulose chromatography and isoelectric focusing.Total activity of acid phosphatase did not change greatly ona fresh-weight basis during maturation and aging. Extracts of immature green tomato (Lycopersicon esculentum Mill.)fruit showed six bands on polyacrylamide gels. Isoelectric focusingand subsequent electrophoresis of the fractions obtained atdifferent pH values, produced evidence for seven acid phosphatasecomponents in tomato extracts. Unlike the situation with certainother climacteric fruit, acid phosphatase in tomato extractsdecreased steadily during ripening to a level of activity only25–40% that of immature green fruit extracts. It is thereforeconcluded that the onset of senescence in climacteric fruitsis not dependent on an increase in acid phosphatase. Substratespecificity, response to inhibitors, Km, and pH optima of partiallypurified fractions were similar to those of tea-leaf extracts. (Received November 29, 1972; )     

Kinetic analysis of the effect of (-)-epigallocatechin gallate on the DNA scission induced by Fe(II)

The DNA strand scission induced by Fe(II) in a citrate buffer solution and the effect of (-)-epigallocatechin gallate (EGCg) were kinetically analyzed. The rate of consumption of dissolved oxygen by Fe(II) in each of these solutions was measured and paralleled that DNA scission. Coordinated EGCg accelerated these reactions. Curves of the time-course characteristics of DNA scission were simulated by using the rate constant of oxygen consumption and by assuming that scission with the hydroxyl radical (OH), which was formed from the dissolved oxygen, proceeded competitively with the scavenging of OH by citrate, Cl- ions and EGCg added. Free EGCg acted as a DNA scission inhibitor to scavenge OH, in contrast to the case of the coordinated one. This analysis is useful for estimating the rate constant of the reaction between an antioxidant and OH, and might provide a new method for measuring the OH-scavenging activity.     

Ciliary Neurotrophic Factor (CNTF) Genotypes and CNTF Contents in Human Sciatic Nerves as Measured by a Sensitive Enzyme-Linked Immunoassay

Abstract: To study the level of ciliary neurotrophic factor (CNTF) in human nervous tissues, we developed a sensitive enzyme-linked immunoassay using a specific antibody against human CNTF. This method allowed us to detect as little as 0.3 ng/ml of human CNTF with good linearity and accuracy. Using this method, CNTF levels were determined in human sciatic nerves obtained at autopsy from 21 amyotrophic lateral sclerosis (ALS) patients and 48 subjects who had died of other neurological diseases. CNTF genotypes were also determined. The results indicated that CNTF levels were high in the normal homozygotes and approximately halved in the heterozygote subjects. There was, however, no significant difference in CNTF levels in the sciatic nerves between ALS and other neurological disease patients, indicating that the CNTF level was mainly determined by its genotypes and that the level in the sciatic nerves was not reduced in ALS patients.     

Pyramidodinium atrofuscum gen. et sp. nov. (Dinophyceae), a new marine sand-dwelling coccoid dinoflagellate from tropical waters

A new marine sand‐dwelling coccoid dinoflagellate Pyramidodinium atrofuscum Horiguchi et Sukigara gen. et sp. nov. is described from Jellyfish Lake, Republic of Palau. The dinoflagellate alternates a non‐motile vegetative stage with a motile gymnodinioid stage within its life cycle. The non‐motile stage is dominant in the life cycle and the dinoflagellate reproduces itself by means of the production of two motile cells. The released motile cell swims only for a short period and is directly transformed into the non‐motile cell. The non‐motile cell is sessile, pyramidal in shape, with a single longitudinal ridge and a double transverse ridge. The surface of the cell wall is covered with many processes. The motile cell has a Gymnodinium‐like morphology, but no apical groove is present. An ultrastructural study revealed that the dinoflagellate possesses typical dinoflagellate organelles. Based on the unique morphology of the vegetative non‐motile stage, we propose a new genus Pyramidodinium for this dinoflagellate, with the type species Pyramidodinium atrofuscum Horiguchi et Sukigara, gen. et sp. nov.