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L A Sherman

Publications and source records attributed to L A Sherman.

At least 73 records · Page 4Linked to original sources
PubMed1994

MapA, an iron-regulated, cytoplasmic membrane protein in the cyanobacterium Synechococcus sp. strain PCC7942.

Growth of Synechococcus sp. strain PCC 7942 in iron-deficient media leads to the accumulation of an approximately 34-kDa protein. The gene encoding this protein, mapA (membrane-associated protein A), has been cloned and sequenced (GenBank accession number, L01621). The mapA transcript is not detectable in normally grown cultures but is stably accumulated by cells grown in iron-deficient media. However, the promoter sequence for this gene does not resemble other bacterial iron-regulated promoters described to date. The carboxyl-terminal region of the derived amino acid sequence of MapA resembles bacterial proteins involved in iron acquisition, whereas the amino-terminal end of MapA has a high degree of amino acid identity with the abundant, chloroplast envelope protein E37. An approach employing improved cellular fractionation techniques as well as electron microscopy and immunocytochemistry was essential in localizing MapA protein to the cytoplasmic membrane of Synechococcus sp. strain PCC 7942. When these cells were grown under iron-deficient conditions, a significant fraction of MapA could also be localized to the thylakoid membranes.

Amino Acid Sequence↗
PubMed1994

Oscillating behavior of carbohydrate granule formation and dinitrogen fixation in the cyanobacterium Cyanothece sp. strain ATCC 51142.

It has been shown that some aerobic, unicellular, diazotrophic cyanobacteria temporally separate photosynthetic O2 evolution and oxygen-sensitive N2 fixation. Cyanothece sp. ATCC strain 51142 is an aerobic, unicellular, diazotrophic cyanobacterium that fixes N2 during discrete periods of its cell cycle. When the bacteria are maintained under diurnal light-dark cycles, N2 fixation occurs in the dark. Similar cycling is observed in continuous light, implicating a circadian rhythm. Under N2-fixing conditions, large inclusion granules form between the thylakoid membranes. Maximum granulation, as observed by electron microscopy, occurs before the onset of N2 fixation, and the granules decrease in number during the period of N2 fixation. The granules can be purified from cell homogenates by differential centrifugation. Biochemical analyses of the granules indicate that these structures are primarily carbohydrate, with some protein. Further analyses of the carbohydrate have shown that it is a glucose polymer with some characteristics of glycogen. It is proposed that N2 fixation is driven by energy and reducing power stored in these inclusion granules. Cyanothece sp. strain ATCC 51142 represents an excellent experimental organism for the study of the protective mechanisms of nitrogenase, metabolic events in cyanobacteria under normal and stress conditions, the partitioning of resources between growth and storage, and biological rhythms.

Carbohydrate Metabolism↗
PubMed1994

Transfusion therapy in solid organ transplantation.

Numerous transfusion issues arise in the support of solid organ transplantation. ABO and HLA can cause graft rejection immunity or problems in transfusion support. Blood usage has been reduced by the use of erythropoietin in renal failure and aprotinin during liver transplants. Multifactorial coagulopathy during liver transplant surgery continues to be a challenge requiring close monitoring, however. The beneficial effect of transfusions on graft survival has lessened in recent years, but remains an area of active research.

ABO Blood-Group System↗
PubMed1993

Alloantibodies can discriminate class I major histocompatibility complex molecules associated with various endogenous peptides.

Molecules encoded by a single major histocompatibility complex class I gene can associate with any one of a large number of peptide ligands. T-cell receptors have the capacity to discriminate among these peptide-class I complexes and in many cases bind only a single peptide-class I complex with sufficient affinity to trigger effector function. In contrast, it is generally assumed that class I-specific alloantibodies are indifferent to peptide heterogeneity, being directed toward allele-specific determinants on the molecule. In this report, three monoclonal antibodies were used to precipitate Kb molecules from cell lysates. Surprisingly, in each case a different set of peptides was found to be associated with Kb as detected by peptide-dependent Kb-specific alloreactive cytolytic T lymphocytes or by biochemical resolution. These results demonstrate that the affinity of binding by alloantibodies can be affected by the endogenous peptide ligand.

Animals↗
PubMed1993

The highly abundant chlorophyll-protein complex of iron-deficient Synechococcus sp. PCC7942 (CP43') is encoded by the isiA gene.

A chlorophyll (Chl)-protein complex designated CPVI-4 becomes the major pigment-protein complex in Synechococcus sp. PCC7942 cells grown under conditions of iron limitation. Work by Laudenbach et al. (J Bacteriol [1988] 170: 5018-5026) has identified an iron-repressible operon, designated isiAB, containing the flavodoxin gene and a gene predicted to encode a Chl-binding protein resembling CP43 of photosystem II. To test the hypothesis that the CP43-like protein is a component of the CPVI-4 complex, we have inactivated the isiAB operon in Synechococcus sp. PCC7942 using directed insertional mutagenesis. Mutant cells grown under conditions of iron limitation exhibit pronounced changes in their spectroscopic and photosynthetic properties relative to similarly grown wild-type cells. Notably, the strong 77 K fluorescence emission at 685 nm, which dominates the spectrum of iron-deficient wild-type cells, is dramatically reduced in the mutant. The loss of this emission appears to unmask the otherwise obscured photosystem II emissions at 685 and 695 nm. Most importantly, mildly denaturing gel electrophoresis shows that mutant cells no longer express the CPVI-4 complex, indicating that the isiA gene encodes a component of this abundant Chl-protein complex.

Bacterial Proteins↗
PubMed1993

Unicellular, aerobic nitrogen-fixing cyanobacteria of the genus Cyanothece.

Two marine, unicellular aerobic nitrogen-fixing cyanobacteria, Cyanothece strain BH63 and Cyanothece strain BH68, were isolated from the intertidal sands of the Texas Gulf coast in enrichment conditions designed to favor rapid growth. By cell morphology, ultrastructure, a GC content of 40%, and aerobic nitrogen fixation ability, these strains were assigned to the genus Cyanothece. These strains can use molecular nitrogen as the sole nitrogen source and are capable of photoheterotrophic growth in the presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea and glycerol. The strains demonstrated a doubling time of 10 to 14 h in the presence of nitrate and 16 to 20 h under nitrogen-fixing conditions. Rapid growth of nitrogen-fixing cultures can be obtained in continuous light even when the cultures are continuously shaken or bubbled with air. Under 12-h alternating light and dark cycles, the aerobic nitrogenase activity was confined to the dark phase. The typical rates of aerobic nitrogenase activity in Cyanothece strains BH63 and BH68 were 1,140 and 1,097 nmol of C2H2 reduced per mg (dry weight) per h, respectively, and nitrogenase activity was stimulated twofold by light. Ultrastructural observations revealed that numerous inclusion granules formed between the photosynthetic membranes in cells grown under nitrogen-fixing conditions. These Cyanothece strains posses many characteristics that make them particularly attractive for a detailed analysis of the interaction of nitrogen fixation and photosynthesis in an aerobic diazotroph.

Bacteria, Aerobic↗
PubMed1993

The molecular basis of allorecognition.

Until recently it was generally assumed that the focus of T cell receptor recognition of allogeneic MHC molecules was the polymorphic regions on the molecule that differed between responder and stimulator. It is now clear that all T cell recognition, including self-tolerance and allorecognition, involves both the MHC molecule and its associated peptide ligand. Polymorphic residues located within the peptide binding groove of the MHC and inaccessible to the T cell receptor can profoundly affect selection and recognition of bound peptides. These peptide differences between histoincompatible individuals greatly amplify the antigenic impact of MHC polymorphism and result in the high frequency of alloreactive cells. Evidence for the role of peptides in allorecognition is reviewed.

Animals↗
PubMed1992

Selecting T cell receptors with high affinity for self-MHC by decreasing the contribution of CD8.

Selective events during T cell repertoire development in the thymus include both the positive selection of cells whose receptors recognize self-major histocompatibility complex (MHC) molecules and negative selection (tolerance) of cells whose interaction with self-MHC is of high affinity. The affinity of T cell interactions with class I MHC molecules includes contributions by both the T cell receptor and the CD8 coreceptor. Therefore, by decreasing the affinity of the interaction with CD8, T cells whose receptors have relatively high affinities for self-MHC may survive negative selection. Such T cells were generated and those T cells reactive with self-MHC plus antigen also displayed low affinity for self.

Animals↗
PubMed1992

Oxygen yield and thermoluminescence characteristics of a cyanobacterium lacking the manganese-stabilizing protein of photosystem II.

Previous experiments have shown that a Synechocystis sp. PCC 6803 mutant (delta psbO) lacking the extrinsic manganese-stabilizing protein (MSP) exhibits impaired, but significant levels of H2O-splitting activity [Burnap, R., & Sherman, L.A. (1991) Biochemistry 30, 440-446]. [14C]DCMU-binding experiments now show that the number and affinity of DCMU-binding sites (normalized to chlorophyll) are equivalent in delta psbO and the wild type, suggesting equal concentrations of assembled reaction centers. A similar conclusion is reached on the basis of measurements of PSII electron transport (DPC-supported DCPIP reduction) by mutant and wild-type thylakoids. The pattern of flash O2 yield by delta psbO cells measured with a bare platinum electrode exhibits a period four oscillation (with a maximum on the third flash), indicating that the H2O-splitting enzyme in delta psbO retains the basic mechanistic features found in normal cells. However, the amplitude of these signals is smaller and more highly damped than those obtained from wild-type cells, suggesting the absence of MSP results in a higher miss probability and/or a reduction in the number of centers competent in oxygen evolution. Analysis of the rise kinetics of the ampermeric signal on the bare platinum electrode indicates that the S3-[S4]-S0 transition is retarded by at least a factor of 5 in the mutant. Thermoluminescence emission peak temperatures indicate that the S2QA-, S2QB-, and S3QB-charge pairs are significantly more stable with respect to recombination in the mutant. The intensities of the thermoluminescence emissions are also significantly reduced in the mutant.(ABSTRACT TRUNCATED AT 250 WORDS)

Binding Sites↗
PubMed1992

Extracellular processing of peptide antigens that bind class I major histocompatibility molecules.

One problem associated with the use of synthetic peptides as antigens in vivo is their susceptibility to inactivation by proteolytic degradation. A situation is described in which a serum protease, angiotensin-converting enzyme (ACE), is actually responsible for the class I binding activity of a commonly used influenza antigen, nucleoprotein (NP)(147-158R-). This peptide has been reported to be a highly efficient class I antigen. Evidence is presented that demonstrates that the peptide is inactive until cleaved by ACE, which is a normal constituent of serum. The enzyme removes a COOH-terminal dipeptide resulting in the sequence NP(147-155), which is identical to the naturally processed peptide. Such extracellular processing of peptides and proteins may occur for a variety of antigens both in vitro and in vivo, and could have important implications for the design of proteolytically resistant vaccines.

Animals↗
PubMed1992

Peripheral tolerance to an islet cell-specific hemagglutinin transgene affects both CD4+ and CD8+ T cells.

To study the basis for immunological tolerance of peripheral tissue-specific antigens, a transgenic mouse line was established that expresses the influenza hemagglutinin (HA) on pancreatic islet beta cells (Ins-HA transgenic mice). When followed up to 14 months of age, Ins-HA transgenic mice did not develop spontaneous autoimmune disease. Upon immunization with HA-expressing viruses, high titers of HA-specific circulating antibody were detected; however, T cell responses by both the T helper and T cytolytic compartment were markedly reduced as compared with transgene-negative littermates, and no evidence could be found for islet infiltrates. Adoptive transfer of histocompatible lymphocytes from transgene-negative mice plus virus into irradiated Ins-HA hosts resulted in islet inflammation dominated by CD4+ T cells, indicating that the HA antigen was accessible to activated T cells. These results suggest that T cells can be rendered tolerant of antigens expressed outside the thymus.

Amino Acid Sequence↗
PubMed1991

Alloreactive T cells discriminate among a diverse set of endogenous peptides.

Previous studies have demonstrated that class I major histocompatibility complex (MHC) molecules are occupied by peptides of endogenously synthesized self proteins. Since graft rejection appears to be mediated by the normal occurrence of high frequencies of cytolytic T lymphocytes (CTLs) specific for allogeneic MHC molecules, it is important to know if such CTLs recognize specific MHC-peptide complexes (as opposed to the MHC molecule per se) and, if so, whether allorecognition is the result of the recognition of a limited spectrum of antigenic determinants or, alternatively, the recognition of a diverse array of MHC-self peptide complexes. This issue has been examined using a mutant cell line, T2Kb, that expresses class I molecules devoid of endogenously derived peptides. This cell line was not recognized by Kb-specific alloreactive CTLs. However, upon exposure to peptides derived by cyanogen bromide cleavage of cytoplasmic proteins these cells became sensitized for recognition and lysis by a majority of the CTL clones examined. Reverse-phase HPLC fractionation of the heterogeneous cell-derived peptides revealed that individual CTL clones were specific for different peptide antigen(s). Thus, the high frequency of alloreactive T cells that is responsible for graft rejection appears to represent the sum of numerous T-cell clones specific for a diverse array of endogenous peptide antigens presented in the context of allogeneic class I molecules.

Animals↗
PubMed1991

Analysis of the HLA-restricted influenza-specific cytotoxic T lymphocyte response in transgenic mice carrying a chimeric human-mouse class I major histocompatibility complex.

Transgenic murine lines have been constructed that express a chimeric class I molecule composed of the alpha 1 and alpha 2 domains of HLA-A2.1 and the alpha 3, transmembrane, and cytoplasmic domains of H-2Kb. Upon immunization with influenza virus, transgenic mice developed a strong A2.1Kb-restricted cytotoxic T lymphocyte (CTL) response specific for the same matrix protein epitope that serves as the dominant A2.1-restricted determinant in the equivalent human response. Fine specificity analysis of CTL clones using truncated peptides revealed strong similarity between the response repertoire of transgenic mice and that previously reported using influenza-specific A2.1-restricted CTL clones from humans. This suggests that even when considering T cell responses by different species, the alpha 1 and alpha 2 domains of the restriction element play a dominant role in determining the CTL specific repertoire. Thus, substituting the alpha 3 domain of A2.1 with a murine counterpart has permitted development of a transgenic strain that should serve as an excellent model system in studies of HLA-restricted responses.

Animals↗
PubMed1991

Deletion mutagenesis in Synechocystis sp. PCC6803 indicates that the Mn-stabilizing protein of photosystem II is not essential for O2 evolution.

The photosystem II (PSII) reaction center complex coordinates a cluster of Mn atoms that are involved in the accumulation of oxidizing equivalents generated by light-induced charge separations within the intrinsic portion of the PSII complex. A 33-kDa extrinsic protein, termed the Mn-stabilizing protein (MSP), has been implicated in the stabilization of two of the four Mn atoms of the cluster, yet the precise role of this protein in O2 evolution remains to be elucidated. Here we describe the construction of a mutant of the cyanobacterium Synechocystis sp. PCC6803 in which the entire gene encoding MSP has been deleted. Northern and immunoblot analyses indicate that other PSII proteins are expressed and accumulated, despite the absence of MSP. Fluorescence emission spectra at 77 K indicate PSII assembles in the mutant, but that the binding of MSP is required for the normal fluorescence characteristics of the PSII complex, and suggest a specific interaction between MSP and CP47. Fluorescence induction measurements indicate a reduced rate of forward electron transport to the primary electron donor, P680, in the mutant. It is concluded that in contrast to previous reports, MSP is not required for the assembly of active PSII complexes nor is it essential for H2O-splitting activity in vivo.

Blotting, Northern↗
PubMed1991

Cell-type-specific recognition of allogeneic cells by alloreactive cytotoxic T cells: a consequence of peptide-dependent allorecognition.

Recently, we provided the first direct evidence that the determinant recognized by some alloreactive cytotoxic T lymphocytes (CTL) clones included an endogenous peptide antigen(s), which could be derived by cyanogen bromide cleavage of cytoplasmic proteins. The studies outlined in this report provide three important findings which confirm and extend our previous results. First, the description of peptide-dependent Kb-specific alloreactive CTL clones is now extended to clones isolated from several responders primed with allogeneic EL4 tumor cells. This indicates that the previously reported results were not unique to the original EL4-primed responder. Second, we establish that peptide-dependent Kb-specific alloreactive CTL clones can be identified after priming with normal allogeneic spleen cells. Thus, induction of peptide-dependent CTL clones is not a unique property of EL4 tumor cells. These results suggest that peptide plays an important role in formation of the allogeneic determinants recognized during graft rejection. Third, these studies reveal that a number of alloreactive CTL clones exhibit cell-type-specific recognition of allogeneic murine target cells. This is consistent with the possibility that some CTL clones recognize peptide antigens that are limited in their tissue distribution. The mechanisms and implications of these findings are discussed.

Animals↗
PubMed1991

Exogenous beta 2-microglobulin is required for antigenic peptide binding to isolated class I major histocompatibility complex molecules.

Binding of antigenic peptides to purified class I major histocompatibility complex (MHC) molecules, as measured by antigen-specific cytolytic T lymphocyte (CTL) degranulation, was found to occur in the presence of serum but not in its absence. The role of soluble beta 2-microglobulin (beta 2m), a normal component of serum, in class I-peptide complex formation was therefore examined. Sera depleted of beta 2m did not support effective peptide binding to class I, but binding was restored in the presence of low concentrations of purified human beta 2m. Sequential incubation of immobilized class I with human beta 2m first, followed by peptide, resulted in antigenic complex formation, while reversing the order of pulsing could not. Similar results were obtained in experiments examining H-2Db, Kb and Kd with appropriate peptides and CTL. These results demonstrate that mature class I proteins are not able to directly bind peptide, but that interaction with exogenous beta 2m results in a structure that will subsequently bind peptide. Binding of exogenous beta 2m appears to result in "empty" class I molecules, possibly by exchange for endogenous beta 2m, with a concomitant loss of endogenous peptide.

Animals↗