Stig R. Erlander

Interface and meniscus widths and positions for low-speed ultracentrifuge runs

Abstract Three No. 4 rubber stoppers placed between the drive base plate and chamber top of a Spinco model E centrifuge gave an extremely steady rotor at speeds around 1,000 rev./min. The broadening of the meniscus and of the interface of two liquid phases at low speeds is thought to be due to the diffraction of light out of the optical system from a curved surface. The entire surface appears to be curved since the meniscus and interface bands broaden both towards and away from the center of rotation as speed is reduced. The position of the meniscus (or the interface) appears to be at a distance of approximately one-third of the meniscus width from the apex of the curved surface. The width of the interfacial band can be reduced by decreasing the interfacial tension between the two liquid phases. The composition of the cell and the shape of the cell sector also appear to alter the width of the meniscus and interfacial bands.

Starch Biosynthesis. 3: The Glycogen Precursor Mechanism Using Phosphorylase in the Production of the Precursor Glycogen

Based on the previously proposed glycogen precursor mechanism (S. R. Erlander, Enzymologia 19 (1958), 273–283), it is now proposed that phosphorylase is the primary enzyme for the production of linear chains in the precursor glycogen. The mechanism involves the translocation of ADPglucose (ADPGlu), and is suppressed by ATP because of a reverse of the ADPglucose pyrophosphorylase (ADPG pp) mechanism. Soluble starch synthase II (SSS II) is a back-up system, involves the translocation of glucose-6-phosphate (Glu-6-P), and is activated, not suppressed, by ATP, and is used if phosphorylase or the ADPGlu translocator is destroyed. Each system is independent and produces products which suppress the other. Hence, only one system works at a time. Both mechanisms produce a glycogen precursor and both are dependent upon ADPGlu pp. The initial higher radioactivity of amylose and the constant yield of amylose can be explained by a three or four day biosynthesis of this glycogen, followed by the removal of the glycogens exterior branches by debranching enzymes to produce amylose and amylopectin. These removed branches are first degraded (using SSS I and II) to ADPGlu, which is the only source of ADPGlu for amylose synthesis. The retention of the polymodal behavior of debranched amylopectin in going from Bomi to shx barley amylopectins is most likely due to a change from phosphorylase to SSS II since both debranched amylopectins produce Poisson distributions.

Starch biosynthesis. II. The statistical model for amylopectin and its precursor plant glycogen

A modified Meyer model as well as a model containing all possible randomly branched structures (a statistical model) were used for both a precursor glycogen and its amylopectin and were modified by placing short A-chains into the interior part of the structure and thus eliminating some of the longer chains. Debranching of the exterior A-chains of the glycogen model produced amylopectin plus 25% amylose with the assumption that only those A-chains removed from the glycogen were converted into amylose. A comparison of the debranched amylopectin model with debranched wheat and barley amylopectins showed the polymodal behavior of debranched amylopectins can be duplicated with a model system that has inner A-chains and a Poisson size distribution. Branching of external chains of glycogen without chain extension after synthesis has stopped plus inner A-chains can explain the increase (from glycogen to amylopectin) in the A/B chain ratio, the existance of cluster structures. and the observed decrease in branching (8% to 4%) in going from dent, waxy and sweet corn glycogens to their respective amylopectins (or 6.3% to 2.0% for glycogen to amylopectin in ae corn). Disperseable (as in waxy) or indispersable high molecular weight aggregates may be due to different amylopectin-protein complexes.

Starch Biosynthesis. I. The Size Distributions of Amylose and Amylopectin and Their Relationships to the Biosynthesis of Starch

Equations for the size distributions of both linear and branched polymers were applied to debranched amylopectin, linear amylose. and branched amylose polymers. The experimental size distribution of linear amylose corresponds to the broad size distribution of an A-B condensation polymer, whereas that of debranched amylopectin linear chains corresponds to the much narrower Poisson size distribution. These dramatic differences illustrate that different types of enzymes synthesize the linear chains of amylose as compared to those of amylopectin. These results support the previously proposed mechanism. The polymodal behavior of debranched amylopectin is due to the existance of individual Poisson-type polymers created by tier structures in a statistically formed precursor glycogen. Equations were developed which enable the calculation of the percentages of these individual Poisson polymers. When applied to the differences between shx and Bomi barley amylopectins, it is concluded that both studies agree that two different short, inner tier, A-chains exist, where the longer chain is located in the more external third tier in the amylopectins. In amylose, three different polymers exist: A linear amylose A-B type condensation polymer, a branched amylose which behaves as a statistical A-R-B 2 type polymer, and an intermediate, non-statistically branched amylose polymer.

Behaviour of polyelectrolytes in ultracentrifugal molecular weight determinations. II. The concentration coefficient B

An approximate form can be assumed for the concentration coefficient B in ultracentrifugal molecular weight determinations. By employing the electrostatic charge obtained from titration studies, this approximate form of the B coefficient predicts a value which is considerably higher than the experimental value for bovine serum albumin (BSA). This discrepancy suggests that the electrical potential term cannot be zero and, hence, that BSA must possess permanent or induced polarization or both. A dipole moment of approximately 1400 DEBYE units was calculated for BSA by employing into the B coefficient a simplified model given by EDSALL and WYMAN. This compares favorably with a dipole moment of 760 units obtained from the dielectric constant of BSA. The (dCr/rdr) vs. Cr curves for BSA were examined for solutions containing a small amount of salt plus acid or for solutions containing the acid alone. When only the acid is present (hydrochloric or trichloroacetic), the concentration coefficient Br appears to decrease as the cell radius r increases. This decrease in Br with r is accompanied by an increase in the total coefficient B which is a function of the initial polymer concentration. The most plausible explanation for this decrease in Br with r is that the hydronium ion concentration decreases with r due to greater binding of the acid anion at the meniscus. This change in pH with r decreases the value of BSAs electrostatic charge Z with cell radius. Thus the value of Br decreases with r because of a corresponding decrease in the positive term — (Z/M)(d ln CB/d CP) and an increase in the absolute value of the negative dipole moment term. The latter term decreases with an increase in r because of a corresponding increase in the number of zwitterions on BSA. Calculations show that the total coefficient B will be increased three- or fourfold for a difference in charge of (Zb−Za) = −0.04, because the coefficient of d(Z′/M′)/dCP is also negative. A small amount of salt (0.0005 molar) eliminates this decrease in Br and increase in B. Bei Molekulargewichtsbestimmungen mit der Ultrazentrifuge kann der Konzentrationskoeffizient B naherungsweise berechnet werden. Unter Verwendung der durch Titration erhaltenen elektrostatischen Ladung ergibt diese Naherung einen B-Wert, der fur Rinder-serumalbumin (BSA) erheblich hoher ist als der gemessene Wert. Dieser Unterschied legt nahe, das der elektrische Potentialterm nicht Null sein kann und daher bei BSA permanente oder induzierte Polarisation oder beides auftritt. Aus dem B-Wert wurde mit einem von EDSALL und WYMANN angegebenen, vereinfachten Modell ein Dipolmoment von etwa 1400 DEBYE fur BSA berechnet. Das entspricht etwa dem Dipolmoment von 760 DEBYE, das sich aus der Dielektrizitatskonstante fur BSA ergibt. Fur saure BSA-Losungen mit und ohne Zusatz einer kleinen Menge Salz wurde (dCr/rdr) gegen Cr aufgetragen. In Gegenwart von Salz- oder Trichloressigsaure allein scheint der Konzentrationskoeffizient Br mit zunehmendem Abstand vom Rotationszentrum abzunehmen. Diese Abnahme von Br mit r ist von einer Zunahme des Gesamtkoeffizienten B, der von der anfanglichen Polymerkonzentration abhangt, begleitet. Die plausibelste Erklarung fur diese Abnahme von Br mit r ist eine Abnahme der H-Ionenkonzentration mit r infolge der starkeren Bindung des Saureanions am Meniskus. Diese pH-Anderung vermindert den Wert der elektrostatischen Ladung von BSA. Also nimmt der Wert von Br ab infolge einer entsprechenden Abnahme des positiven Terms −(Z/M) (dln CB/dCP) und einer Zunahme des Absolutwertes des negativen Dipolterms. Der zweite Term nimmt ab infolge einer entsprechenden Zunahme der Zahl der Zwitterionen auf dem BSA-Molekul. Fur eine Ladungsdifferenz Zb−Za = −0,04 ergibt die Rechnung eine drei- oder vierfache Zunahme des Gesamtkoeffizienten B, weil der Koeffizient von d(Z′/M′)/dCP auch negativ ist. Bei Zusatz von wenig Salz (0,0005 molar) verschwindet diese Abnahme von Br und die Zunahme von B.