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D P Delmer

Publications and source records attributed to D P Delmer.

12 recordsLinked to original sources
PubMed1995

A membrane-associated form of sucrose synthase and its potential role in synthesis of cellulose and callose in plants.

Sucrose synthase (SuSy; EC 2.4.1.13; sucrose + UDP reversible UDPglucose + fructose) has always been studied as a cytoplasmic enzyme in plant cells where it serves to degrade sucrose and provide carbon for respiration and synthesis of cell wall polysaccharides and starch. We report here that at least half of the total SuSy of developing cotton fibers (Gossypium hirsutum) is tightly associated with the plasma membrane. Therefore, this form of SuSy might serve to channel carbon directly from sucrose to cellulose and/or callose synthases in the plasma membrane. By using detached and permeabilized cotton fibers, we show that carbon from sucrose can be converted at high rates to both cellulose and callose. Synthesis of cellulose or callose is favored by addition of EGTA or calcium and cellobiose, respectively. These findings contrast with the traditional observation that when UDPglucose is used as substrate in vitro, callose is the major product synthesized. Immunolocalization studies show that SuSy can be localized at the fiber surface in patterns consistent with the deposition of cellulose or callose. Thus, these results support a model in which SuSy exists in a complex with the beta-glucan synthases and serves to channel carbon from sucrose to glucan.

Amino Acid Sequence↗
PubMed1995

Genes encoding small GTP-binding proteins analogous to mammalian rac are preferentially expressed in developing cotton fibers.

In animals, the small GTP-binding proteins, Rac and Rho, of the ras superfamily participate in the signal transduction pathway that regulates the organization of the actin cytoskeleton. We report here on the characterization of two distinct cDNA clones isolated from a cotton fiber cDNA library that code for homologs of animal Rac proteins. Using gene-specific probes, we have determined that amphidiploid cotton contains two genes that code for each of the two Rac proteins, designated Rac13 and Rac9, respectively. The gene for Rac13 shows highly enhanced expression in developing cotton fibers, with maximal expression occurring at the time of transition between primary and secondary wall synthesis. This is also the time at which reorganization of the cytoskeleton occurs, and thus the pattern of expression of Rac13 is consistent with its possible role, analogous to animal Rac, in the signal transduction pathway that controls cytoskeletal organization.

Amino Acid Sequence↗
PubMed1991

beta-Furfuryl-beta-glucoside. An endogenous activator of higher plant UDP-glucose: (1----3)-beta-glucan synthase.

We have recently established the existence of endogenous activators of higher plant UDP-glucose: (1----3)-beta-glucan synthase (Callaghan, T., Ross, P., Weinberger-Ohana, P., and Benziman, M. (1988) Plant Physiol. 86, 1099-1103). Here we report the purification and chemical analysis of the most abundant and specific compound, termed Activator I, isolated from Vigna radiata. This compound was extensively purified by a multistep procedure which yielded 0.1 mg of purified activator/g of fresh tissue. Enzyme digestion, neutral sugar analysis, GC/MS of permethylated derivatives, and NMR analysis of native Activator I indicated that the compound contains a single beta-linked glucosyl residue. High resolution FAB-MS indicated an elemental composition of C11H16O7 (Mr = 260), with a calculated Mr of 98 for the aglycone. 13C, DEPT, and COSY NMR spectra showed that the aglycone molecule is an oxygen heterocycle of 5 carbons, consistent with a structure of beta-furfuryl alcohol. Comparison of IR and GC/EI-MS spectra of authentic beta-furfuryl alcohol with native aglycone confirmed the conclusion that Activator I is beta-furfuryl-beta-glucoside. Chemically synthesized beta-furfuryl-beta-glucoside has identical chemical properties and biological activity when compared with the purified endogenous activator (Ka = 50 microM).

Enzyme Activation↗
PubMed1985

New approaches to the study of cellulose biosynthesis.

Examples of a variety of approaches for studying the mechanism and regulation of cellulose biosynthesis are presented. Attempts to demonstrate conclusively a cellulose synthase activity using membrane preparations derived from higher plants have not been successful; the predominant UDP-glucose: beta-glucan-beta-glucosyltransferase detected in these preparations is a beta-(1----3)-glucan synthase that is dependent upon Ca2+ and a beta-glucobiose, such as laminaribiose or cellobiose, for activity. Xyloglucan glucosyltransferase activity is detected in all plant preparations examined and, in cotton fibres, the activity of this enzyme correlates well with the level of xyloglucan found in the fibre. Low activity for a Mg2+-dependent beta-(1----4)-glucan synthase is found in extracts from soybeans and mung beans, but not cotton fibres; this enzyme could represent either a dissociated form of xyloglucan glucosyltransferase or a partially latent form of cellulose synthase. In vitro translation of RNA from developing cotton fibres shows notable increases in the level of several relatively abundant translatable mRNAs associated with the time of onset of secondary-wall cellulose synthesis. In order to determine whether these apparent changes in gene expression represent enhanced expression of specific genes required for cellulose synthesis, several strategies are being developed for identification of specific polypeptides required for this process. One strategy involves the successful development of a technique for detection of glucan synthase activity in acrylamide gels, a technique that should prove useful for characterization of the polypeptide composition of such enzymes. We have also synthesized a photoaffinity analogue of 2,6-dichlorobenzonitrile (DCB), a potent and specific inhibitor of cellulose synthesis. The analogue is also an effective inhibitor in vivo, and upon ultraviolet irradiation of extracts in the presence of the radioactive analogue, we observe a relatively specific labelling of a polypeptide of 18 X 10(3)Mr. Finally, we have studied the spatial regulation and structural requirements for cellulose synthesis in internode cells of the alga Chara corallina. Cellulose deposition in vivo shows spatial localization, which correlates with acid and base bands along the cell. Using internodes perfused with solutions containing UDP-[14C]glucose and subsequently ligated, we were able to demonstrate synthesis of a highly insoluble cell-wall-localized glucan, thus offering hope that Chara can be developed as another useful system for studying the mechanism and regulation of cellulose synthesis.

Cell Membrane↗
PubMed1982

Achievement of high rates of in vitro synthesis of 1,4-beta-D-glucan: activation by cooperative interaction of the Acetobacter xylinum enzyme system with GTP, polyethylene glycol, and a protein factor.

Regulatory properties of a cellulose synthase (UDP-forming)(UDPglucose:1,4-beta-D-glucan 4-beta-D-glucosyltransferase, EC 2.4.1.12) have been demonstrated by using enzyme preparations derived from cells of Acetobacter xylinum. Preparation of a particulate fraction in the presence of 20% (wt/vol) polyethylene glycol-4000 (PEG-4000) yields enzyme with activity 3- to 10-fold higher than that previously reported. The enzyme prepared in this fashion also shows a further marked, specific activation by GTP. The Ka for GTP is 34 microM. Guanosine 5'-[gamma-thio]triphosphate, an analog of GTP, is even more effective than GTP (Ka for guanosine 5'-[gamma-thio]triphosphate = 17 microM). A large number of other nucleotides and nucleotide derivatives were tested with no effect. Enzyme prepared in the absence of PEG-4000 does not respond to GTP because it lacks a protein factor necessary for GTP activation. PEG-4000 promotes the interaction of the protein factor with the enzyme. The factor itself has no synthase activity nor does it stimulate activity of the enzyme in the absence of GTP. In the presence of GTP, protein factor, and PEG-4000, initial rates of enzyme activity 200 times greater than those previously reported can be achieved. Such rates exceed 40% of the in vivo rate of cellulose synthesis from glucose.

Bacterial Proteins↗
PubMed1975

Circadian rhythms in Neurospora crassa: oscillation in the level of an adenine nucleotide.

In a mutant strain (bd) of Neurospora, the biological clock is visibly expressed at the growing front of a mycelial mat by sequential periods of conidiating (spore-forming) and non-conidiating growth. The edges (8 mm) of the mycelium at different ages were sampled during a 31 h period, and the adenine nucleotide levels were enzymatically assayed. In the edge region, the total adenosine 5'-monophosphate (AMP) level showed an oscillation, with a minimum of 0.5 mumol/g (residual dry weight) and a maximum of 6.0 mumol/g. The total adenosine 5'-triphosphate level and the total adenosine 5'-diphosphate level showed no obvious oscillation. The oscillation in AMP content had many of the properties of a circadian rhythm. Its period was about 22 h long, it was phase-shifted by light, and it was damped out by continuous illumination. The oscillation in AMP level led to an oscillation in the overall cellular energy charge from 0.65 to 0.93. However, the energy charge calculation does not take into account any possible compartmentalization of AMP, and therefore must be interpretated cautiously. It is suggested that the underlying cause of the oscillation in AMP level could be a rhythmic, partial uncoupling of mitochondrial oxidative phosphorylation.

Adenine Nucleotides↗