Lower Apparent Molecular Weight Research Articles

Expression of structurally diverse Qa-2-encoded molecules on the surface of cloned cytotoxic T lymphocytes.

Extracts of 125I-labeled cloned murine cytotoxic T lymphocytes (CTL) were immunoprecipitated with alloantisera to the cloned CTL and rabbit antisera to beta-2 microglobulin. Polyacrylamide gel electrophoresis (PAGE) of the specific precipitates revealed, as expected, 125I-labeled components that corresponded to products of class I genes of the major histocompatibility complex (MHC). However, additional class I gene products of relatively low apparent molecular weight (Mr) were also observed. Similar analyses of spleen cells from a variety of MHC-congenic mouse strains suggested that the class I molecules of relatively low Mr are encoded in the Qa-2 region of the MHC, and this was confirmed by immunoprecipitation with a monoclonal antibody to Qa-2. Surprisingly, however, the cell surface Qa-2 molecules of different CTL clones differed in Mr, in isoelectric focusing (IEF) pattern, and in the number of distinguishable molecules expressed per clone: some clones seemed to express only a single Qa-2-encoded molecule while others expressed two distinct ones. Treatment of the immunoprecipitated Qa-2 with endoglycosidase F (Endo F) resulted in a decrease in Mr of approximately 5,000-6,000, corresponding to the expected loss of N-linked oligosaccharides, but the decrease did not eliminate structural variability among the clones. Structural diversity of the Qa-2-encoded molecules expressed on CTL could arise because CTL clones differ (a) in the particular Qa-2 genes they express, (b) in the way they splice Qa-2 gene transcripts or, perhaps, (c) in Endo F-resistant oligosaccharides on their Qa-2 molecules.

Characterization and purification of a soluble protein controlling Ca-channel activity in paramecium

The analysis of the voltage-sensitive Ca ++ channel of the unicellular eucaryote, Paramecium has been extended to a biochemical level based on recent observations that the transfer of cytoplasm from wild-type cells into mutants lacking Ca ++-channel function (“pawn” in P. tetraurelia and “CNR” in P. caudatum) causes mutant cells to regain Ca ++-channel function. We have microinjected various cytoplasmic fractions into mutant cells and measured the restored Ca ++-channel function using a convenient behavioral assay. Following the “curing” activity, we characterized and purified the component from wild-type cytoplasm that can restore the function missing in cells carrying mutations in the cnrC gene. The curing factor is not an RNA, but a heat-labile, -SH-containing protein that appears to affect existing mutant channels on the ciliary membrane. We have purified this factor over 500-fold from the soluble cytoplasm using conventional techniques. The protein is of low apparent molecular weight (<30,000 daltons), acidic, soluble, and does not have the properties of calmodulin.

Biochemical characteristics of the heterophile transplantation antigen (HT-A).

The Heterophile Transplantation Antigen (HT-A) is a clinically important antigen which is found in some human kidneys and on the erythrocytes and in the serum of rats and some other mammals. Recent reports suggest the possibility that the anti-HT-A antibodies responsible for graft rejection may cross react with one or more HL-A specificities. The ideal way to investigate this possibility would be to perform careful serological and biochemical comparisons of purified HT-A and HL-A. We report here the first stage of these investigations, the examination of the biochemical properties of HT-A. The HT-A molecule of rat plasma is apparently a glycoprotein with a molecular weight greater than 1,000,000. The HT-A antigenic determinant site seems to reside in the carbohydrate portion of the glycoprotein. The native HT-A molecule is apparently susceptible to proteolytic enzymes of rat plasma and serum and is cleaved by these enzymes to produce a fragment with a lower apparent molecular weight and a larger fragment which coelutes with lipoproteins during Sepharose CL-4B chromatography. The data presented here should allow development of a rational purification scheme for the HT-A glycoprotein of rat plasma to be used in further studies in the relationship between HT-A and HL-A.

Integrity of glycoprotein complex sugars is required for homing but not for several other membrane‐mediated functions

In order to correlate the biochemistry of cell surface carbohydrates with cell function, we have treated cells with swainsonine and followed the biochemical and functional modifications induced by this compound. After treatment with swainsonine, surface glycoproteins had a lower apparent molecular weight and a higher isoelectric point. This is compatible with the replacement of complex carbohydrates by hybrid or high-mannose carbohydrates. Several functional tests were unaffected. However, swainsonine-treated cells displayed an altered pattern of in vivo homing, suggesting that carbohydrates play a role in this process.

Interaction of platinum with metallothionein-like ligands in the rat kidney after administration of cis-dichlorodiammine platinum II

After the administration of the anticancer drug cis-dichlorodiammine platinum II (cisplatin) to male rats, the Pt in the soluble fraction of the kidney is isolated, by gel filtration, in association with a high molecular weight component and a low molecular weight fraction. At 24 h, Pt is also recovered in a metallothionein-like fraction which elutes from Sephadex G-50 with a lower apparent molecular weight than endogenous (Cu, Zn)-thionein or Cd-thionein isolated from the kidneys of Cd 2+-treated rats. None of these low molecular weight metal-binding fractions binds to Octyl Sepharose CL-4B. On DE-52 ion exchange chromatography, Cd-thionein is resolved into two isometallothioneins whereas the low molecular weight Pt-binding fraction is only partially purified and contains at least six components which elute at higher gradient concentrations than metallothionein. Pretreatment with Cd 2+ which stimulates the synthesis of renal and hepatic metallothionein has no effect on the uptake and subcellular distribution of Pt in the liver and kidneys. Cisplatin treatment reduces the concentration of Cu and Zn in the renal metallothionein and other soluble protein fractions in the kidney. When administered to Cd 2+-pretreated rats, cisplatin promotes the loss of Zn from the soluble protein fractions but causes the redistribution of Cd from the metallothionein to the high molecular weight fraction and fails to inhibit the Cd 2+-induced accumulation of Cu in the kidneys and the binding of Cu to the soluble protein fractions. It is suggested that metallothionein probably does not have a significant role in the renal metabolism of Pt following the administration of cisplatin to rats.

Identification of la Antigens on the Murine Adenocarcinoma L T-85&lt;xref ref-type="fn" rid="FN2"&gt;2&lt;/xref&gt;&lt;xref ref-type="fn" rid="FN3"&gt;3&lt;/xref&gt;&lt;xref ref-type="fn" rid="FN4"&gt;4&lt;/xref&gt;

Antigens associated with the H-2Kk and I-Ak regions of the major histocompatibility complex have been identified with monoclonal antibodies on an in vivo grown murine alveologenic adenocarcinoma, LT-85. Immunoprecipitation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis demonstrated differences in the molecular structure of I-Ak region-coded antigens between LT-85 and host C3H/HeN mammary tumor virus-negative (MTV-) or autologous C3HfB/HeN splenocytes. Ia antigens derived from LT-85 tumor cells exhibited both an increased heterogeneity in the alpha-chain and a lower apparent molecular weight in the beta-chain. Tryptic peptide mapping of the I-Ak antigen alpha- and beta-chains derived from C3H/HeN MTV- mice and LT-85 tumor cells revealed a single peptide difference in the beta-chains of these antigens. Results obtained with neuraminidase-treated I-Ak beta-chains indicated that this difference was not due to sialic and content. Maintenance of LT-85 in vitro, even for short periods, resulted in the loss of these I-Ak antigens. However, this loss of I-Ak antigen expression was fully reversible with in vivo growth.

Effect of swainsonine on the processing of the asparagine-linked carbohydrate chains of alpha 1-antitrypsin in rat hepatocytes. Evidence for the formation of hybrid oligosaccharides.

The biosynthesis of the proteinase inhibitor alpha 1-antitrypsin has been studied in rat hepatocyte primary cultures. Newly synthesized alpha 1-antitrypsin was found in hepatocytes as a glycoprotein of an apparent molecular weight of 49,000 carrying oligosaccharide side chains of the high mannose type. In the hepatocyte medium a secreted alpha 1-antitrypsin of an apparent molecular weight of 54,000 could be identified as a glycoprotein with carbohydrate chains of the complex type. Pulse-chase experiments revealed a precursor-product relationship for the two forms of alpha 1-antitrypsin. When the hepatocytes were treated with swainsonine, an intracellular form of alpha 1-antitrypsin with an apparent molecular weight of 49,000 indistinguishable from that of control cells was found. However, the alpha 1-antitrypsin secreted from swainsonine-treated hepatocytes was different from that present in control media. It was characterized by a lower apparent molecular weight (51,000), a higher amount of [3H]mannose incorporation, half as much incorporation of [3H]galactose, and the same amount of [3H]fucose incorporation compared to alpha 1-antitrypsin of control media. In contrast to the 54,000 complex type alpha 1-antitrypsin from control media the 51,000 alpha 1-antitrypsin from the medium of swainsonine-treated cells was found to be susceptible to the action of endoglucosaminidase H, even when fucose was attached to the proximal GlcNAc residue. alpha 1-Antitrypsin secreted from swainsonine-treated cells combines features usually associated with either high mannose or complex type oligosaccharides and therefore represents a hybrid structure. In spite of its effect on the carbohydrate part of alpha 1-antitrypsin swainsonine did not impair the secretion of the incompletely processed glycoprotein.

Cell-free synthesis of phytochrome apoprotein

A single polypeptide is immunospecifically precipitated by monospecific antiphytochrome from the total translation products of both wheat-germ and rabbit-reticulocyte cell-free protein synthesizing systems programmed with oat (Avena sativa L.) poly(A) RNA. The mobility of this polypeptide is slightly lower on sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis than that of immunoaffinity-purified, 118 kdalton phytochrome and corresponds to an apparent molecular weight of 124 kdalton. Evidence against the possibility that this mobility difference results from intracellular processing of the 124-kdalton protein is provided by extraction of freeze-dried tissue directly into boiling SDS-containing buffer. This procedure yields a phytochrome species with a mobility on SDS polyacrylamide gel electrophoresis indistinguishable from that of the in-vitro translation product. Together the data indicate that the phytochrome polypeptide is synthesized in its mature form in the cell but is subject to modification to a form with lower apparent molecular weight during immunopurification.

Influence of dissolved humic materials on carbon assimilation and alkaline phosphatase activity in natural algal‐bacterial assemblages

SUMMARY. Mixed natural assemblages of algae and bacteria exhibited lower rates of 14C assimilation and high rates of dissimilation of recent photosynthate when amended with low concentrations (7.2 mg 1‐1) of unfractioned dissolved humic materials (DHM). The extent of the inhibition or stimulation was greatest in the smaller (1–5 μm) assemblage particles. In different algal‐bacterial assemblages, additions of DHM markedly enhanced community alkaline phosphatase activity (APA), particularly under low light regimes, DHM of low apparent molecular weight was much more stimulatory to both 14C assimilation and APA than DHM of high apparent molecular weight, supporting the belief that DHM molecular weight is an important determinant of DHM interactive capacity. Higher concentrations of D HM (either unfractionated, or molecular weight fractionated) produced greater APA responses. Addition of phosphate enhanced the disparity in rates of 14C assimilation of samples incubated under low and high light regimes, increased the rates of 14C assimilation, and depressed APA. There were indications of interactions between DHM and phosphorus in several experiments. Two hypotheses were invoked to explain increases in APA in response to DHM: (1) increased competition between algae and bacteria for phosphate following bacterial release from substrate limitation, or (2) DHM may have acted as a sequestering agent for organophosphorus compounds, and in so doing, gradually depleted available phosphate. In either case, it is clear that DHM alters phosphorus cycling. This DHM characteristic may be ecologically as important as its ability to complex trace metals.

The purification, characterization, and activation of phosphoenolpyruvate carboxykinase from chicken liver mitochondria.

The enzyme phosphoenolpyruvate carboxykinase has been purified from chicken liver mitochondria. This purification includes a pseudo-affinity column step utilizing Sepharose 4B-blue dextran which binds the enzyme. The enzyme elutes with ITP to yield protein which is greater than 98% pure. The enzyme has Mr = 75,400 +/- 200 estimated by high speed sedimentation equilibrium and 70,500 +/- 500 estimated by reduced sodium dodecyl sulfate-polyacrylamide gels. The enzyme is abnormally retarded on molecular exclusion resins yielding low apparent molecular weight values. The amino acid analysis indicates that the enzyme has a high proline content and a high tryptophan content and contains 9 mol of cysteine/mol of enzyme. No disulfide bonds were detected. The extinction coefficient (epsilon 1% 280 = 16.5 +/- 0.1) reflects the high tryptophan content. The Svedberg coefficient (s20,w = 4.63 +/- 0.03 S) is consistent with a globular protein of Mr = 70,500-75,400. The activation of the enzyme was investigated by steady state kinetics. The carboxylation reaction has an activation energy of 17.6 kcal/mol. There is no requirement of a monovalent cation for activity. A thiol is necessary for maximal activity, although apparently not to reduce disulfide bonds within the enzyme. Incubation with dithiothreitol stabilizes enzymatic activity but beta-mercaptoethanol facilitates loss of activity. The kinetics of activation by Mn2+ was performed. The Ks value for phosphoenolpyruvate (300 microM) decreases to an apparent Km of 67 microM with increasing concentrations of Mn2+. The concentration of Mn2+ does not affect the interaction of HCO-3 with the enzyme, however. Analysis of data in terms of free IDP indicates that increasing Mn2+ decreases the Km of IDP but analysis as MnIDP indicates the Km,app of MnIDP is independent of the Mn2+ concentration. The enzyme interacts with Mn2+ with a KA = 67 microM and the Km,app decreases to a value of 8 microM with saturating substrates. The substrate analogue (Z)-3-fluorophosphoenolpyruvate is a good substrate for the reaction (Km = 30 microM) with 27% Vmax compared to P-enolpyruvate (Km = 180 microM). Except for 3-bromophosphoenolpyruvate, other analogues have shown weak competitive or noncompetitive inhibition. Potential analogues of oxalacetate (succinate, citrate, isocitrate, malate, and alpha-ketoglutarate) all elicit weak (greater than 15 mM) inhibition.

Hypersecretion of an Abnormal Form of Follicle-Stimulating Hormone Associated with Suppressed Luteinizing Hormone Secretion in a Woman with a Pituitary Adenoma*

Pituitary tumors producing FSH have hitherto been reported only in males, all of whom have had normal or raised LH levels in serum. This report describes a female with a pituitary adenoma associated with supranormal serum levels of FSH. The FSH was also qualitatively abnormal when compared with FSH in the serum of other postmenopausal women, had a lower apparent molecular weight on gel chromatography, and was less negatively charged, as shown by electrophoresis. The results of LRH tests and suppression tests with ethinyl estradiol indicated autonomy of the FSH-producing adenoma. The FSH level increased concomitant with tumor enlargement and decreased after surgical removal of the pituitary adenoma or pituitary irradiation. The serum level of the glycoprotein alpha-subunit was raised about 100-fold. Any free FSH beta-subunits were not detectable in serum. The abnormal FSH had antigenic sites in common with both the alpha- and beta-subunits of FSH. The LH level was extremely low, and there was no response to LRH tests or ethinyl estradiol treatment. After gel chromatography, a small amount of LH, corresponding to 1/50th of the average for the patient's age, was detected at the position for normal LH. There was no GH response to insulin-induced hypoglycemia, while the cortisol increase was normal. Thyroid and adrenal functions were normal. The PRL level was within the normal range and increased slightly after estrogen treatment.

Thermostable, ammonium-activated malic enzyme of Clostridium thermocellum

‘Malic’ enzyme ( l-malate:NADP + oxidoreductase (oxaloacetate-decarboxylating, EC 1.1.1.40) was purified from Clostridium thermocellum by DEAE-cellulose, agarose-NADP and Sephadex G-200 column chromatography. The 117-fold purified ‘malic’ enzyme displayed a maximum activity of 135 units/mg at 40°C and represented 0.8% of the total cell protein. Sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis analysis of the protein suggested 90% purity and an approximate tetrameric subunit molecular weight of 40 000. The enzyme absolutely required both bivalent and monovalent cations for catalysis. Mn 2+ and NH 4 + were the most effective cationic activators examined. Increasing NH 4 + concentration increased both enzyme activity and affinity toward l-malate. The apparent K m for l-malate was 3 · 10 −4 M at 0.4 mM NH 4Cl. Enzyme activity increased linearly when temperature was raised between 22–60°C and a Q 10 of 2.1 was calculated from an Arrhenius plot. The enzyme was stable to heating at 60°C but was denatured at higher temperatures. The enzyme half-life was 10 min at 72°C. The enzyme displayed a broad pH optimum (7.2–8.2 for Tris-HCl buffer) but was inactivated by p-chloromercuribenzoate. The high thermal stability, low apparent molecular weight and NH 4 + activation are properties not common to all previously described ‘malic’ enzymes.

Cell cycle-dependent expression of surface antigens in murine T-lymphoma cells: II. Murine leukemia virus gp70 increases in S phase

The surface expression of the murine leukemia virus (MuLV) major envelope protein, MuLV gp70, is shown here to be cell cycle-dependent. AKR mouse T-lymphoma cells, BW 5147, synchronized by double thymidine blockade, were collected in S or G1 phase and lactoperoxidaselabeled with 125I and 131I. The iodinated surface proteins were solubilized with 0.05% Triton X-100 and aliquots of each preparation containing equal amounts of protein were mixed and precipitated by goat anti-Rauscher gp70 (70000 D MuLV envelope glycoprotein) serum and Staphylococcus aureus or by rabbit anti AKR gp70 serum and sheep anti-rabbit Ig serum. The precipitates were solubilized, reduced and separated in 9% SDS-PAGE gels. The gels were sliced and analyzed in a gamma-counter programmed for double-labeled samples. The gp70 molecule, precipitated by both antisera had the same quantitative cell cycle variability and the same pI heterogeneity as the 70 K antigen precipitated by an antiserum made against BW 5147 cell membranes at the G1 phase of the cell cycle. The expression of MuLV gp70 on BW 5147 cells is increased 2-fold in the S phase of the cell cycle as compared to the G1 and G2 phases. This increase is associated with one of the two gp70 components which has a lower isoelectric point and a lower apparent molecular weight.

Routes of thrombin action in the production of proteolytically modified, secondary forms of antithrombin-thrombin complex.

The reaction between thrombin and antithrombin results in the formation of an inactive, stable, equimolar complex between the two proteins. However, under most reaction conditions several secondary complex forms, which have lower apparent molecular weights in dodecyl sulfate/polyacrylamide gel electrophoresis, appear concomitantly with or immediately following the production of the primary form of the complex. Purification of nascent, intact complex and treatment of this complex form with thrombin demonstrated that these subsidiary forms of antithrombin-thrombin complex may arise by proteolysis of the nascent complex by excess thrombin. Dissociation of such proteolytically modified complex preparations by hydroxylamine, and examination of the dissociation products by dodecyl sulfate/polyacrylamide gel electrophoresis suggested that degradation occurs primarily in the thrombin part of the complex, and only after prolonged proteolysis in its antithrombin moiety also. Incubation of antithrombin with several autolytically modified thrombin preparations showed that formation of subsidiary complex forms can also occur by an alternative route, i.e. between premodified thrombin forms and the inhibitor. In contrast, complex formation between thrombin and active forms of antithrombin, which have been modified by thrombin before complex formation, is unlikely, since no such active forms of antithrombin could be demonstrated.

Activation in vitro of rheumatoid synovial collagenase from cell cultures.

Rheumatoid synovial cells dissociated from matrix and adherent to culture dishes released a latent form of collagenase into culture medium. Previous studies have shown that the latent enzyme does not complex with alpha2-macroglobulin and binds to fibrillar substrate. We now show that serum-free culture medium of the synovial cells contains an inhibitor of collagenase as well as latent enzyme; the two were separated on a column of acrylamide/agarose. Latent collagenase (estimated mol wt 45,000-49,000) was transformed by trypsin to active collagenase of approximately equal to mol wt 33,000. When mixed with inhibitor the active enzyme formed an inactive complex again with approximately equal to mol wt 45,000-49,000. The inhibitor(s) itself was found in one major peak of mol wt 33,000-35,000 and several minor peaks eluting with lower apparent molecular weight. Mersalyl, an organic mercurial compound, effectively activated latent collagenase producing an active enzyme with approximately equal to mol wt 33,000. Bacterial collagenase did not activate latent enzyme. We suggest that latent rheumatoid synovial collagenase, as it is harvested from synovial cells in culture, is an enzyme-inhibitor complex.

The genome-linked protein of picornaviruses IV. Difference in the VPg's of encephalomyocarditis virus and poliovirus as evidence that the genome-linked proteins are virus-coded

A protein (EMC-VPg) covalently attached to EMC RNA has been found whose properties are distinct from those of the genome-linked protein of poliovirus (polio-VPg). EMC-VPg isolated from virus grown in HeLa cells is identical to EMC-VPg from virus grown in L cells, as determined by urea sodium dodecyl sulfate (SDS)/polyacrylamide gel electrophoresis. In contrast, polio-VPg isolated from virus grown in HeLa cells migrates more rapidly in these gels indicating a lower apparent molecular weight. These data show that the physical properties of the genome-linked proteins of two picornaviruses depend upon the nature of the viral genome rather than what proteins are available in the host cells. The most straightforward explanation for this phenomenon is that VPg is virus-coded. Evidence is presented which suggests that, as in poliovirus, VPg is attached to the EMC genome at the 5′ terminus via the nucleotide pUp.

Cell-surface changes in Ricinus communis toxin (ricin)-resistant variant of a murine lymphoma.

A variant of the murine lymphoma cell line BW5147 that was 250 times more resistant than the parent to Ricinus communis II agglutinin (RCAII, ricin) toxicity (measured in the absence of serum) was selected by repeated exposure of cells to increasing concentrations of the lectin. Quantitative binding of the lectin, however, was decreased by only 30-40% in the variant. In contrast with several reported lectin-resistant variants, most surface glycoproteins on the parental and variant cell surfaces were similar, as judged by electrophoresis after lactoperoxidase-catalyzed iodination and RCAI-affinity chromatography. Surface studies showed that an RCAI- and RCAII-binding protein of about 80,000 daltons on the surfaces of parental cells is altered on the variant cells to a form with a lower apparent molecular weight. We suggested that this protein is important for entry of RCAII molecules in parental cells, but that its altered form on the variant cells no longer mediates efficient RCAII uptake, thus imparting toxin resistance. In addition, a protein of approximately 35,000 daltons, which does not bind RCAI, is weakly lactoperoxidase-iodinated on parental but not variant cells.

Macromolecular distribution near the limits of density-gradient columns. Some applications to the separation and fractionation of glycoproteins

1. Expressions are derived for the distribution at density-gradient equilibrium of macromolecules whose densities are (a) close to the values characterizing the solution limits or (b) outside the span of the gradient. 2. Density-distribution predicted by the expressions agree with those obtained by rigorous methods. 3. The distribution equations are applied to hypothetical mixtures of proteins and glycoproteins in commonly used density-gradient media to simulate separation and fractionation conditions. 4. It is shown that CsBr, although less efficient than CsCl for fractionation, is nevertheless adequate for most purposes; in analytical experiments it may often have advantages over CsCl. Limitations on the use of LiBr are explored. 5. An expression is derived which allows the variance of the partial specific volume of the macromolecular component to be determined from the variance of the buoyant density. It is shown that the relative resolving powers of different salts is expressed by their values of the quantity (formula: see text). 6. The equations are applied to a well-characterized glycoprotein preparation at equilibrium in CsCl and in Cs2SO4:it is shown that the much wider distribution in CsCl than in Cs2SO4 is explicable in terms of the variance in buoyant density and the solvation properties of the salts. 7. Limitations of the expressions arise when dispersity in density is represented by a low apparent molecular weight; realistic simulations can then only be obtained when the component is fully banded.

The influence of physicochemical factors on the thermal inactivation of murine interferon.

The degradation of biological activity of virus-induced murine interferon was determined in linear nonisothermal and multiple isothermal tests. The stabilizing effect of pH during heating on interferon in solution was greatest at low pH, such that pH 2 greater than pH 5 greater than pH 7 greater than or equal to pH 9; freeze-dried preparations of interferon were also more heat-stable at acid pH than at neutral pH. Heat stability was a function of the H+-ion concentration rather than the ionic composition of the buffer; interferon solutions containing monovalent cations with different ionic radii had similar heat stability. A change in the H+ ion concentration was a critical event during the cooling of heated interferon: a shift in the direction of acidity contributed to stability whereas a shift towards alkalinity led to inactivation. The rate of cooling of heated interferon significantly influenced its residual activity. Rapid cooling and sudden freezing decreased the residual activities of interferons at pH 2 and 9 more than "normal" cooling, an effect not observed at pH 7. Interferon heated to 80degree C could not be reactivated at 40degree C or 55degree C. Interferon of higher apparent molecular weight was more heat-stable than that with lower apparent molecular weight. It is postulated that the physicochemical alterations in the aqueous environment significantly affecting the stability of interferon operate by producing changes in the size and/or conformation of interferon molecules. A model is proposed that relates thermal inactivation to different possible molecular states of interferon.

Sequential polypeptides. VI. The synthesis of some sequential polypeptide collagen models containing proline analogues.

Syntheses of poly-(L-alanylglycyl-L-thiazolidine-4-carboxylic acid) and of poly-(L-piperidine-2-carbonyl-L-alanylglycine)via the corresponding tripeptide pentachlorophenyl esters and of poly-(glycyl-L-piperidine-2-carbonyl-L-alanine) and poly-(glycyl-L-azetidine-2-carbonyl-L-alanine)via the corresponding tripeptide 2-hydroxy-phenyl esters are described. All of the polydisperse preparations were of low apparent molecular weight except the polymer containing thiazolidine-4-carboxylic acid, which on viscosity criteria had a weight average molecular weight in the region of 10.000.