SYNTHESIS, PURIFICATION AND STRUCTURE ACTIVITY RELATIONSHIPS IN THE DODECAPEPTIDE MATING FACTOR OF SACCHAROMYCES CEREVISIAE.
Item
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Title
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SYNTHESIS, PURIFICATION AND STRUCTURE ACTIVITY RELATIONSHIPS IN THE DODECAPEPTIDE MATING FACTOR OF SACCHAROMYCES CEREVISIAE.
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Identifier
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AAI8401915
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identifier
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8401915
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Creator
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BAFFI, ROBERT ANGELO.
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Contributor
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Fred Naider
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Date
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1983
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Language
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English
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Publisher
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City University of New York.
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Subject
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Biology, Microbiology
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Abstract
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The (alpha)-factor is a tridecapeptide, Trp-His-Trp-Leu-Gln-Leu-Lys-Pro-Gly-Gln-Pro-Met-Tyr, which mediates the sexual conjugation of a and (alpha) haploids of Saccharomyces cerevisiae. Five desTrp('1)-dodecapeptide analogs were prepared in which Trp('3) was replaced with (beta)-cyclohexylalanine (Cha) while Leu('6) was substituted by Ala, Val, Ile, Nle or D-Leu. Also synthesized was desTrp('1), Phe('3)-dodecapeptide. All peptides were prepared by solution phase techniques utilizing mixed anhydrides or HOBT active esters. Purification to > 98% homogeneity was accomplished by reverse-phase HPLC.;Structure-function relationships for positions 3, 6, 7, 9 and 12 of the des Trp('1)-dodecapeptides were determined using quantitative bioassays which monitor aberrant cell morphologies (shmoos), increased agglutination, or cell cycle arrest of a-cells. In addition to the 6 peptides whose preparation is reported herein, 18 other dodecapeptides (14, 16, B. J. Kundu unpublished results) were investigated. Replacement of Trp('3) with Ala or Phe led to inactive analogs which were antagonists for biological activities. Replacement of Trp('3) with Cha enhanced morphogenic activity 3-fold while decreasing agglutination activity 5-fold. Elimination of the charge on Lys('7) led to a 3 to 11-fold decrease in morphogenic potency and a 40-fold decrease in agglutination potency. Substitution of Leu('6) with either Nle, Ile or Val caused a decrease in morphogenic activity of 1 to 10-fold while agglutination potency decreased from 35 to 375-fold. Replacement of Leu('6) with Ala or D-Leu resulted in inactive analogs which were not antagonists for (alpha)-factor activity. These results indicate that size, hydrophobicity and configuration influence the activity of position six analogs.;Substitution of Gly('9) with D-Ala led to a slight increase in morphogenic potency while the activity of the L-Ala('9) analog was reduced 48-fold. Replacement of Met('12) with Nle resulted in a peptide of equal morphogenic activity.;Competition studies with inactive position 3 analogs demonstrated that agglutination activity could be inhibited with lower concentrations of antagonists than were required to inhibit morphogenic activity. Furthermore, the ratio of morphogenic activity to agglutination activity varied nearly 500-fold for the active analogs. These results indicate a heterogeneous response of a-cells to (alpha)-factor and suggest that a-cells contain more than 1 receptor for (alpha)-factor.
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Type
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dissertation
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Source
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PQT Legacy CUNY.xlsx
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degree
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Ph.D.
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Program
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Biochemistry
