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Biomedical subjects

A Spector

Publications and source records attributed to A Spector.

At least 73 records · Page 4Linked to original sources
PubMed1979

Further investigation of the role of calcium in human lens protein aggregation.

High-molecular-weight (HMW) protein from human cataractous lenses, isolated by differential centrifugation, was deaggregated in 7M urea and then reaggregated in either the presence or absence of 10 mM CaCl2. Over 90% of the material reaggregated in the presence of calcium appears to have a size greater than 50 X 10(6) daltons. By contrast, only 20% to 25% of the material reaggregated in the absence of calcium has molecular weight greater than 50 X 10(6) daltons. Disulfide formation during reaggregation is unlikely in the latter experiment, since the addition of 50 mM mercaptoethanol caused no change in results. About 60% to 70% of the low-molecular-weight (LMW) protein fraction deaggregated in 7M urea buffer can be converted to HMW species in the presence of 10 mM CaCl2, when the deaggregating agent is removed. However, only 5% to 10% of this protein is converted to HMW species if the deaggregation step is eliminated. Experiments with 45 Ca indicate that whereas calcium is necessary for the formation of the HMW aggregates, only one calcium per approximately 5 X 10(5) daltons remains bound in the reaggregated material. The data suggest that although calcium may be required to induce aggregation to HMW species, it is not required to stabilize such macromolecules. SDS-polyacrylamide gel electrophoresis of the HMW species formed upon reaggregation of the dissociated HMW species with calcium indicates the presence of all the major polypeptide subunits of the original HMW species present in the lens; however, reaggregation in the absence of calcium yields HMW species lacking in the 9600 dalton component.

Calcium Chloride↗
PubMed1978

Detection of bityrosine in cataractous human lens protein.

Bityrosine was isolated from the insoluble protein of human cataractous lenses. Identification was based on correspondence with synthetic bityrosine with respect to chromatography, fluorescence, and ultraviolet and mass spectra. It is suggested that the compound may form cross-links with polypeptide chains in old and cataractous lenses, causing significant alteration in native protein structure.

Cataract↗
PubMed1978

Disulfide-linked high molecular weight protein associated with human cataract.

A major high molecular weight disulfide-linked protein has been isolated from cataractous lenses. It is only present in the water-insoluble protein fractions. This species has not been found in normal lenses of comparable age. Upon reduction of this fraction, polypeptides having molecular weights of approximately 60,000, 43,000, and 20,000 as well as a noncharacterized heterogeneous species are released. Similar sized polypeptides have been found in various noncovalently linked aggregates in both normal and cataractous lenses. Examination of the disulfide-linked protein fraction indicates that approximately 70% of the sulfhydryl groups are in the oxidized state. Although little change in the sizes of the other major polypeptides in the water-insoluble fraction is observed upon reduction, these components were also found to contain an appreciable disulfide content. Such results indicate that the only major lens fraction containing disulfide-linked polypeptide is the high molecular weight species and that the disulfides present in the remaining fractions are either intrachain disulfides or link polypeptides to small peptides.

Cataract↗
PubMed1978

Racemization in human lens: evidence of rapid insolubilization of specific polypeptides in cataract formation.

After early life, the dry weight of normal human lenses increases at a relatively constant rate with time. Transformation from soluble to insoluble material appears to occur at a comparable rate, resulting in a constant amount of soluble material. However, in cataract the insolubilization rate is accelerated. These observations are supported by determination of D-aspartic acid/L-aspartic acid ratios. The abundance of D-aspartic acid increases with aging at a constant rate in the insoluble fraction of normal lenses but does not change in the soluble fraction. However, in cataractous lenses there is a significant decrease in the ratio in the insoluble fraction. Examination of polypeptides isolated from reduced and alkylated soluble and insoluble cataractous lens proteins as well as other data suggest the following additional conclusions: (i) the 10,000-dalton polypeptide in the insoluble fraction is derived in part from degradation of an already insoluble precursor; and (ii) the lowered abundance of D-aspartic acid in the insoluble fraction of cataractous lenses is primarily due to the rapid insolubilization of the 43,000- and 20,000-dalton range components.

Age Factors↗
PubMed1977

Isolation and characterization of initiation fragments from lens 10S and 14S alpha-crystallin messenger ribonucleic acids.

The alpha-crystallin 10S and 14S messenger ribonucleic acids (mRNAs) for the B and A chains, respectively, were isolated from calf lenses. Initiation complexes were formed with both mRNAs after which the unprotected regions were digested with ribonuclease T1. A single fragment of approximately 45 nucleotides was obtained from both the 10S and 14S mRNAs. The fragments retained the ability to reform initiation complexes under standard conditions. Two-dimensional fractionation of ribonuclease T1 digests indicated considerable similarity between the 10S and 14S fragments. However, marked differences in the (U)G region were observed. The addition of the methylating agent S-adenosyl-L-methionine to the mRNA initiation system increases complex formation form two to five times, suggesting that methylation may be required for initiation.

Animals↗
PubMed1977

The bicistronic nature of lens alpha-crystallin 14S mRNA.

The A2 and B2 polypeptide chains of calf lens alpha-crystallin are synthesized on a 14S, 1500-nucleotide mRNA and a 10S, 735-nucleotide mRNA, respectively. The 10S mRNA is theoretically compatible with the size of the B2 chain, but the 14S mRNA contains approximately twice the required number of nucleotides necessary for A2 chain synthesis. This fact raises the question of the function of the additional nucleotide sequence in the 14S mRNA. The following observations on 14S mRNA suggest that it may contain an additional cistron. (i) Under a number of denaturing conditions, 14S mRNA continues to retain its initial size characteristics. (ii) In addition to synthesis of the A2 chain, 14S mRNA directs the synthesis of another polypeptide with the same electrophoretic mobility as that of the B2 chain. (iii) Molecular hybridization of the 14S mRNA with the cDNA produced from the 10S mRNA suggests that 2 mol of the cDNA bind to 1 mol of the 14S mRNA. (iv) Examination of the nucleotide sequences of the 10S and 14S mRNAs by two-dimensional maps of RNase A and T1 digests indicates marked similarity. The overall data suggest that the additional cistronic component may carry coding information for an alpha-crystallin polypeptide or a closely related polypeptide species.

Animals↗
PubMed1976

Human alpha-crystallin: characterization of the protein isolated from the periphery of cataractous lenses.

alpha-Crystallin has been isolated from the peripheral region of old cataractous lenses. It was found to be closely related to bovine alpha-crystallin and to human newly synthesized alpha-crystallin in terms of its amino acid composition, the size of its polypeptide chains and the lack of free NH2-terminal groups. However, in contrast to the simple urea gel electrophoretic polypeptide patterns obtained with the reference proteins, 11 polypeptides were detected in the preparation. Ten of the polypeptides were isolated and shown to be either A or B chains on the basis of their amino acid compositions and comparison of the peptide maps of their tryptic hydrolysates. The four B chains as well as the six A chains were closely related, with most of the tryptic peptides being common to all members of their respective group. A nomenclature based upon the urea gel electrophoretic mobilites of the polypeptides has been proposed to define each chain. It was found that this alpha-crystallin preparation is composed of at least two populations of macromolecules, one of which contains macromolecules greater than 5 X 10(6) daltons on the basis of gel filtration with Bio-Gel A-5m. The compositions of the two fractions were found to be essentially identical.

Aged↗