David Garfinkel

Studies on pig liver microsomes I. Enzymic and pigment composition of different microsomal fractions.

Abstract 1. 1. A possible seasonal effect in pig liver microsomes is described. 2. 2. A crude fractionation of microsomes into light and bulk fractions is described. Examination in the electron microscope indicates that these contain predominantly smooth-surfaced and rough-surfaced microsomes, respectively. 3. 3. Chemical analysis and observation of the behavior of the microsomes on digestion and solubilization indicate that the smooth- and rough-surfaced microsomes are biochemically distinct, with the smooth-surfaced ones being rich in cytochrome b 5 whereas the rough-surfaced ones are rich in noncytochrome heme and contain a tan pigment. 4. 4. Properties of some microsomal components, including a CO-binding pigment and a soluble yellow pigment, have been described. 5. 5. The binding of hemoglobin to microsomes is described. 6. 6. The effects of low pH and solubilizing agents on microsomes are described. 7. 7. The properties of a microsomal substance (designated “acceptor”) which is reduced by cytochrome b 5 are described, and its origins and importance are discussed.

A Model of Glutamate Metabolism in Brain: A Biochemical Analysis of a Heterogeneous Structure

Glutamate and related amino acids in brain have long attracted the attention of many scientific investigators. The suggested beneficial effects of glutamate on behavior was one of the factors stimulating research on the metabolic aspects of glutamate in brain, a research area already actively pursued in the thirties by Krebs, Weil-Malherbe, and others (Weil-Malherbe, 1950). There is no doubt that Waelsch, through his involvement in the reported beneficial effects of glutamate on epileptic patients, was stimulated to concentrate on the metabolism of glutamate in the brain.

A machine-independent language for the simulation of complex chemical and biochemical systems☆

Abstract A language for the simulation of the kinetic behavior of complex chemical or biochemical systems is described and defined. It is written in FORTRAN IV and should run on any medium or large size batch-processing computer (except that machines of short word-length will require double-precision arithmetic). It permits a chemist to build and manipulate a complex model in his own language, as the input statements are essentially chemical reactions. This language converts these to differential equations, solves them from input boundary conditions which are specified in a manner convenient to the user, and edits the results. Intervention in these processes to meet specialized user requirements is provided for. Applications and possible future developments are discussed.

Magnesium in cardiac energy metabolism

Free intracellular magnesium ion, which influences many metabolic processes, is the subject of ongoing research. Its concentration has been difficult to measure because the available methods, including dye injection, microelectrodes, and nuclear magnetic resonance measurements, are invasive or indirect. Concentrations ranging from 0.1 mM in frog muscle to 6 mM in barnacle muscle have been reported. We describe recent experimental evidence regarding the concentration of free intracellular magnesium and consider the limitations of these methods. A substantial body of evidence, including our models of cardiac energy metabolism and its magnesium-related processes, indicates that intracellular concentrations of free magnesium are low (ca. 0.4 mM) and vary with time and conditions.

A convenient computer program for fitting enzymatic rate laws to steady-state data

Abstract A computer program for the estimation of kinetic parameters in enzymatic rate laws is described. The program includes statistical tests for the reliability of the optimal parameter values and for bias error due to a poor model. A set of criteria for selecting between models is given. The program is easy to learn, convenient to use and modify, inexpensive to use, and widely applicable. The behavior of the program with the correct and an incorrect model was examined using simulated test data. The effects of experimental design and experimental error were investigated.

Isolation and properties of cytochrome b5 from pig liver

Abstract 1. 1. An isolation and purification of cytochrome b 5 from pig liver microsomes is described. Methods are given for obtaining either a purified product (containing 0.31% Fe) or one which also contains DPNH-cytochrome b 5 reductase. 2. 2. A large-scale preparation of microsomes is described. 3. 3. The behavior of the cytochrome on chromatographic columns and in the presence of certain inhibitors, its stability to heat, dialysis, and surface denaturation, and some of its spectrographic properties, both at room temperature and at liquid air temperature, are described. 4. 4. The probable relationship of the isolated cytochrome to the cytochrome as it occurs in microsomes is discussed.

A COMPARATIVE STUDY OF ELECTRON TRANSPORT IN MICROSOMES.

Abstract 1. 1. A comparative study of electron-trasporting components (cytochrome b 5 , flavin, “CO-binding pigment” and total heme) and activities (DPNH-cytochrome c reductase, DPNH-cytochrome b 5 reductase, TPNH-cytochrome c reductase, etc.) has been carried out on microsomes from various vertebrate and invertebrate sources. 2. 2. Microsomal electron-transporting components and activitites are found to be correlated with each other, the pattern of occurence in the microsomes being more characteristic of the organ than of the organism. The order of decreasing concentration and activity of the microsomes is: (by organ), liver, kidney, lung, intestinal mucosa (when functional), testes, adrenal glands, spleen, pancreas and muscle; (by organism), mammals, chicken, turtle, carp, frog, lobster. 3. 3. Of the various functions which have been proposed for cytochrome b 5 , this distribution is most consistent with involvement in steroid hydroxylation and related activities (such as drug metabolism). 4. 4. A previously unknown pigment of nucleated red blood cells (spectroscopically similar to lactoperoxidase) was observed.

Computer simulation of metabolism in palmitate-perfused rat heart. II. Behavior of complete model

Intermediary metabolism in rat hearts persfused with 11 mM glucose plus 1 mM palmitate was simulated by a computer model. Several enzyme submodels in a previous version of the isolated rat heart computer model wre improved, and a new fatty acid oxidation pathway model was added. Compartmentation of metabolites in a pseudostationary state was calculated, and its implications are discussed, e.g., citrate level may not regulate glycolysis because it is mostly mitochondrial. Citrate synthetase, controlled largely by its inhibitors, is of key importance in regulating fatty acid metabolism. The response of aconitase to the mitochondrial Mg2+ levels is of major importance in setting both the mitochondrial citrate and isocitrate levels. Pyruvate dehydrogenase is about 96% in the inactive phosphorylated form, and the active form is also 15% inhibited by products, severely limiting pyruvate oxidation and causing preferential utilization of palmitate as the metabolic fuel. The simulation is consistent with a creatine phosphate shuttle which delivers high energy phosphate to the site of its utilization for mechanical work.

Digital Computer Simulation of an Ecological System, Based on a Modified Mass Action Law

A digital computer simulation of a six—species ecological system is reported. The mass—action (Lotka—Volterra) mechanism is used as a starting point, with modifications as required by ecological reality; the computer program used permits description of the system in terms of species and their interactions, without requiring manipulation of differential equations. The behavior of the simulated system is in good general agreement with reality, and some testable hypothesis have been found. Properties of the system include: the total amount of protoplasm in the system determines the highest level predator it will support; two species cannot simultaneously occupy the same food—chain position; important stabilizing influences include sufficient predator populations and negative feedback controls on reproduction; ecological systems may have distinctive time cycle lengths, resistant to perturbation. See full-text article at JSTOR

Simulation of the detailed regulation of glycolysis in a heart supernatant preparation

Abstract A study of the regulation of the glycolytic pathway has been carried out by simulating in detail the behavior of three glycolyzing beef heart supernatant preparations. Enzyme activity profiles as well as kinetic data on glycolytic intermediates and adenine nucleotides were obtained in the laboratory using glucose-6-phosphate as a substrate. Phosphoglycerate mutase appears to be the limiting enzyme, while triosephosphate isomerase, the most active enzyme when measured separately, is relatively inactive in the complete system. A model consisting of 74 simultaneous differential equations representing 107 chemical reactions were fitted to these results. Incorporated into this model were detailed models of 13 enzymes and crude models of the pentose shunt and ATPase. For most data points, the fit appears fairly good. Most of the enzyme models show strong inhibition as glycolysis proceeds. Individual enzymes were modeled with data from the literature obtained from isolated enzymes. The action of only one enzyme could be satisfactorily simulated without some modification in the form of additional activations, inhibitions, and enzyme-enzyme interactions. This model suggests that the activity of many enzymes is a function of several glycolytic intermediates or products arranged to slow the rate of glycolysis as glycolytic products pile up.