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B L Rogers

Publications and source records attributed to B L Rogers.

At least 37 records · Page 2Linked to original sources
PubMed1993

Immunoaffinity chromatography of recombinant Amb a I in the presence of a denaturing agent.

Recombinant proteins expressed in E. coli are often sequestered into inclusion bodies and require the use of denaturing agents in order to solubilize them. The recombinant form of Amb a I, the major allergen from short ragweed pollen, is one such protein. In some cases solubility can be maintained after the removal of the denaturing agent, particularly if the protein can be folded into its native conformation. However, not all proteins refold readily and after the removal of the denaturing agent the proteins will reaggregate and/or precipitate. In the case of Amb a I, the recombinant protein stays in solution at low concentrations but aggregates with itself and other proteins. The recombinant Amb a I is not expressed at high levels and may be toxic to E. coli. Therefore, isolation from a complex mixture of E. coli proteins was necessary. Monoclonal antibodies which recognize the denatured form of Amb a I were available, allowing for immunoaffinity purification. However, because the protein was not monomeric, this chromatographic technique did not provide an improvement in the purity level when run in normal buffer solutions. Analysis of one monoclonal antibody's stability to urea indicated it could tolerate the presence of 2 M urea and recover full activity. Use of this antibody as an immunoaffinity reagent in a column run in 2 M urea, which minimized aggregation of the E. coli produced proteins, gave a high degree of purification of recombinant Amb a I in one step. This illustrates the potential for the use of denaturing and other solubilizing agents in immunoaffinity chromatography of recombinant proteins.

Allergens↗
PubMed1993

Recombinant Fel d.I: Expression, purification, IgE binding and reaction with cat-allergic human T cells.

This study describes the properties of the two recombinantly expressed polypeptide chains of Fel d I, the major allergen produced by the domestic cat (Felis domesticus). An inframe linker encoding polyhistidine has been added to the 5' ends of the Fel d I chains 1 and 2 cDNAs to facilitate purification using Ni2+ ion affinity chromatography. This method provides high yields in a single step of rchain 1 and rchain 2 of Fel d I with a > 90% level of purity. Polymerase chain reaction (PCR) methods were used to introduce a thrombin cleavage site (LVPR decreases GS) at the N-terminus of both chains. Thrombin cleavage of rchain 1 and rchain 2 followed by HPLC purification of the cleavage products allowed the isolation of each recombinant chain with only two additional residuals (GS) at the N-terminus of the native sequence. Amino acid sequencing analysis of the N-terminus and mass spectrometry of these polypeptides demonstrated that they are highly pure and full-length. Direct ELISA assays showed that IgE from cat-allergic patients binds to both rchain 1 and rchain 2 of Fel d I, demonstrating that both these chains contribute to the allergenicity of this heterodimeric protein. An examination of the reactivity of T cells derived from cat-allergic patients revealed that both polypeptide chains contribute to the T cell response to this allergen. Consequently, it is concluded that the immunological response to Fel d I is composed of a reaction at both the B and T cell level to each of the two chains that constitute the native allergen.

Allergens↗
PubMed1993

Purification and immunochemical characterization of recombinant and native ragweed allergen Amb a II.

The complete sequence of a cDNA encoding Amb a II and its relationship to the Amb a I family of allergens has recently been described [Rogers et al. (1991) J. Immun. 147, 2547-2552; Griffith et al. (1991a), Int. Archs Allergy appl. Immun. 96, 296-304]. In this study, we present results generated with rabbit antipeptide antisera that recognize Amb a II or Amb a I, but not both. The specificity of two anti-Amb a II antipeptide sera, anti-RAE-50.K and anti-RAE-51.K, was verified on Western blots of recombinant Amb a II and Amb aI.1. These two sera, directed against separate regions of the Amb a II molecule, detected three individual 38-kDa Amb a II isoforms on 2D Western blots of aqueous ragweed pollen extract. These Amb a II isoforms have pI in the 5.5-5.85 range and can be easily distinguished from Amb a I isoforms with pI in the 4.5-5.2 range detected by an anti-Amb a I specific peptide antiserum. The Amb a II isoforms have also been individually purified from pollen, positively identified as Amb a II by amino acid sequencing, and visualized as separate bands on IEF gels. An analysis of Amb a II cDNA sequences generated by PCR led to the prediction of three Amb a II isoforms with pI of 5.74, 5.86 and 5.97 that are very similar to the pI deduced from 2D Western blot analysis. Recombinant Amb aI.1 and Amb a II have been expressed in E. coli, purified in their denatured form, and examined by ELISA for their capacity to bind pooled allergic human IgE. Purified native Amb a and Amb a II from pollen were shown to have very similar IgE-binding properties. In contrast, Amb a II had a markedly reduced IgE-binding capacity as compared to Amb a I.1. These data suggest that recombinant Amb a I.1 and Amb a II, isolated in a denatured form, differ significantly in their IgE-binding properties whereas the native molecules isolated from pollen do not.

Allergens↗
PubMed1993

Native and recombinant Fel dI as probes into the relationship of allergen structure to human IgE immunoreactivity.

To delineate the relationship between the structural conformation and the stability of an allergen and its antigenicity, we have chosen the major allergen from cat dander, Fel dI. From protein sequence analysis data we have examined the structure of the naturally occurring Fel dI and we have found it to exist as an anti-parallel heterodimer. We have used ELISA, RAST, Western blot and histamine release techniques to compare the IgE reactivity of a set of cat allergic patient samples to purified, native Fel dI and the E. coli expressed chains 1 and 2. Results from these studies demonstrate a significant level of IgE reactivity to all forms when examined for direct binding. However, both blot and ELISA competition assays show a much higher reactivity to Fel dI in solution compared to the separate recombinant chains and this is supported by the histamine release data. Although native Fel dI chain 2 contains an N-linked carbohydrate moiety, this does not seem to play a role in the reactivity of IgE to chain 2. Denaturation of Fel dI with alkali conditions leads to a dramatic decrease in IgE reactivity, even though measurable changes to the backbone structure of the protein are minimal. One proposed explanation is that both chains possess a core region determined by their primary structures and that the major IgE epitopes are dependent upon them. The relative reactivity amongst these allergen forms varied with the method of analysis, implying that the conformational requirements for IgE antibody binding are best studied by the application of more than one experimental protocol. Results from these qualitative analyses afford insight into the allergenicity of this exceptionally stable cat pelt protein.

Allergens↗
PubMed1993

Children's nutritional status in female-headed households in the Dominican Republic.

A national representative household survey of food consumption, income, and expenditure was conducted in the Dominican Republic in 1986 by Tufts University School of Nutrition in cooperation with USAID Office of Nutrition [1: Rogers B. L. and Swindale A. Determinants of Food Consumption in the Dominican Republic. USAID/S & T/Nutrition, Washington, DC, 1988]. Out of 1440 families surveyed 706 had children under 6 years of age. Anthropometric indicators of height and weight were collected for all 1251 children in the sample in a follow-up study conducted from December 1986 to January 1987 by Tufts with USAID/Santo Domingo Mission funding. Anthropometric measurements were converted to standard deviation scores using NCHS standards; nutritional status was thus measured by height-for-age (HAZ), weight-for-age (WAZ), and weight-for-height (WHZ). Earnings in families with children having males as heads-of-household were roughly one-third greater than in those with females as household head; total income was also greater in these families, though not significantly. More calories and protein per adult equivalent were available in male-headed families as well. In spite of this superior economic and dietary situation, there was a trend throughout the entire sample for children of female-headed households to be taller and heavier for their age than those of two parent homes; all three anthropometric measures showed differences in the same direction. In the lowest expenditure quartile, WAZ and WHZ were significantly greater for children in female-headed households than their counterparts in male-headed households (WAZ: P = 0.01, WHZ: P = 0.00).(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1993

Comparison of the costs of compliance with nutrition education messages to improve the diets of Bangladeshi breastfeeding mothers and weaning-age children.

Local market prices in rural Bangladesh were used to compute the costs of filling the nutrient gaps between actual intakes and safe nutrient requirements, and the costs of compliance with nutrition messages, for 78 lactating mothers and 61 weaning-age breastfed children. (The gap is the difference between the requirement and the amount of nutrient consumed.) To fill the mother's energy gap of approximately 1050 kcal (4393 kJ) would cost an additional 21% of the daily wage, or almost double the value of food she was presently eating. Given social reality, these costs would probably be much greater, as the mother would also need to increase the allocation of food to other household members. The weaning-age children's energy gap could theoretically be closed for less than one-third of the cost of improving the mothers' diets, or about 8% of the daily wage. The increase in food intake equivalent to 2% of the daily wage actually achieved through nutrition education resulted in a significant improvement in child weight gain, though not ideal. These findings suggest that, in the absence of programs which reduce economic barriers, it is economically feasible for families to close the nutrient gaps for weaning-age breastfed children in Bangladesh, but not for lactating women. Thus, education to improve women's diets should be incorporated into programs that make these improvements affordable, whereas education to improve weaning-age children's diets can be implemented with or without other program supports.

Asia↗
PubMed1992

Expression and genomic structure of the genes encoding FdI, the major allergen from the domestic cat.

The genes encoding chain 1 (Ch1) and chain 2 (Ch2) of the major allergen of the domestic cat, Felis domesticus I, have been analyzed by genomic cloning and by polymerase chain reaction (PCR). Ch1 has two potential leader sequences, designated A and B. Analysis of a genomic clone encoding Ch1 demonstrated that one structural gene contains sequences corresponding to both leaders, which utilize different Met start codons. PCR analysis showed that genes encoding Ch1 and Ch2 are co-expressed in both the salivary glands and the skin, and that leader sequence A of Ch1 is utilized preferentially in both tissues. Ch2 was shown to have two dominant forms that are differentially expressed in the aforementioned tissues. The long form (Ch2L), composed of 92 amino acids (aa), is preferentially expressed in the salivary glands, while the short form (Ch2S), composed of 90 aa, is preferentially expressed in the skin. There is minor sequence polymorphism in both forms of Ch2. A genomic clone for Ch2 only contained sequences for Ch2S, suggesting that Ch2L is encoded by an exon not contained within this genomic clone.

Allergens↗
PubMed1992

Evaluation of the impact of weaning food messages on infant feeding practices and child growth in rural Bangladesh.

In rural Bangladesh, a community-based weaning intervention used volunteers to teach complementary feeding to families of 62 breast-fed infants aged 6-12 mo. Over 5 mo, treatment children gained on average 0.46 SD (approximately 460 g) more in weight-for-age (WAZ) than the 55 control subjects, and were approximately 0.5 kg heavier at the final measure. The differences were statistically significant (P < 0.001). The percent median weight-for-age (WAPM) of treatment children held steady at 76% of the National Center for Health Statistics' reference, whereas the WAPM of control subjects dropped from 78% to 72%. The increase in percentage points of severe malnutrition (below -3 WAZ) was only 5% in the treatment group compared with 26% in the control subjects. Treatment children consumed a significantly greater percent of their energy and protein requirements from complementary foods than did control subjects. The affordable complementary foods consisted mainly of cereal porridge with oil and brown sugar. These findings suggest that educational interventions teaching families to feed hygienic, simple, cheap, energy-enriched complementary foods to breast-fed infants after 5-6 mo can improve child growth, even under impoverished conditions.

Bangladesh↗
PubMed1992

Project director characteristics in the Elderly Nutrition Program.

The federally funded Elderly Nutrition Program (ENP) began in 1972, authorized under Title 7 of the Older Americans Act. Its purpose was to serve elders, targeting those in greatest social and economic need. This article describes the characteristics of the Nutrition Project Directors in the ENP through data gathered by a questionnaire completed by a random sample of 430 directors across the nation. One important finding was that directors of programs serving large numbers of low-income or minority elders tended to have less education and commanded lower salaries than directors serving fewer poor and minority elders. In addition, while a higher proportion of these directors were minority members, numbers of nonwhite Nutrition Project Directors remains low.

Educational Status↗
PubMed1991

Amino acid sequence of Fel dI, the major allergen of the domestic cat: protein sequence analysis and cDNA cloning.

The complete primary structure of Fel dI (International Union of Immunological Societies nomenclature), the major allergen produced by the domestic cat, Felis domesticus, was determined by protein sequence analysis and cDNA cloning. Protein sequencing of Fel dI from an immunoaffinity-purified extract of house dust revealed that the allergen is composed of two polypeptide chains. Degenerate oligonucleotides derived from the protein sequence were used in polymerase chain reaction amplification of cat salivary gland cDNA to demonstrate that the two chains are encoded by different genes. Chain 1 of Fel dI shares amino acid homology with rabbit uteroglobin, while chain 2 is a glycoprotein with N-linked oligosaccharides.

Allergens↗
PubMed1991

Complete sequence of the allergen Amb alpha II. Recombinant expression and reactivity with T cells from ragweed allergic patients.

This study defines the complete primary structure of Amb alpha II, an important allergen produced by short ragweed (Ambrosia artemisiifolia). The deduced amino acid sequence derived from the cDNA indicates that Amb alpha II shares approximately 65% sequence identity with the Amb alpha I multigene family of allergens. Full-length cDNA encoding Amb alpha I.1 and Amb alpha II have been expressed in E. coli and purified. An in-frame linker encoding polyhistidine has been added to the 5' end of the cDNA to facilitate purification using Ni2+ ion affinity chromatography, yielding greater than 90% pure recombinant protein in a single step. T cells from patients allergic to ragweed proliferate in response to pollen extract as well as purified recombinant Amb alpha I.1 and Amb alpha II. T cell lines established using either Amb alpha I.1 or II as the stimulating Ag exhibit a high level of cross-reactivity to both proteins. This result is entirely consistent with the extensive primary sequence identity shared by these two proteins. These data suggest that allergic humans recognize shared T cell epitopes on these two related molecules.

Allergens↗
PubMed1991

Multiple Amb a I allergens demonstrate specific reactivity with IgE and T cells from ragweed-allergic patients.

The relationship between the structure and abundance of an inhaled protein and its potential for causing an allergic response is unknown. This study analyzes Amb a I, a family of related proteins formerly known as Ag E, that comprise the major allergens of short ragweed (Ambrosia artemisiifolia). T cells isolated from ragweed allergic patients were shown to proliferate in response to purified Amb a I.1 protein from pollen in in vitro secondary cultures, demonstrating the presence of T cell stimulatory epitopes in Amb a I.1. Three recombinant forms of Amb a I (Amb a I.1, Amb a I.2, and Amb a I.3) obtained as cDNA derived from pollen mRNA were expressed in bacteria. All three recombinant forms were shown to be specifically recognized by pooled ragweed-allergic human IgE on immunoblots, confirming these gene products are important allergens. An examination of immunoblots probed with sera derived from allergic patients revealed a variation in IgE binding specificity. A minority of patients' IgE exclusively reacted with recombinant Amb a I.1, whereas most patients' IgE reacted with Amb a I.1 as well as Amb a I.2 and Amb a I.3 proteins. A detailed examination of the reactivity of T cells derived from 12 allergic patients to these recombinant Amb a I forms revealed that these allergens are all capable of stimulating T cell proliferation in in vitro assays. It is concluded that the allergic response to ragweed pollen in most allergic patients is composed of a reaction to multiple related Amb a I proteins at both the B and T cell levels.

Allergens↗
PubMed1991

Cloning of Amb a I (antigen E), the major allergen family of short ragweed pollen.

To determine the structure of Amb a I (previously called antigen E), the major allergen from short ragweed, cDNA from pollen was cloned into lambda gt11 and lambda gt10. One of the three distinct clones isolated from the lambda gt11 library by screening with anti-denatured Amb a I antibodies was used to screen both libraries for other Amb a I sequences. Multiple clones were isolated and sequenced and proved to be highly homologous but nonidentical. The clones could be divided into three groups based on sequence similarity, and in accordance with the International Union of Immunological Societies-approved nomenclature (Marsh, D. G., Goodfriend, L., King, T. P., Lowenstein, H., and Platts-Mills, T. A. E. (1986) Bull. WHO 64, 767-770) they have been designated Amb a I.1, Amb a I.2, and Amb a I.3. Clones within a group have greater than 99% identity, and similarity among groups is 85-90% at the nucleotide level. The amino acid sequence of four peptides (isolated from antigen E obtained from the Research Resources Branch of the National Institutes of Health) containing 132 amino acids was identical to one of the clones (Amb a I.1). The presence of multiple naturally occurring isoelectric forms of Amb a I was demonstrated by two-dimensional gel electrophoresis and Western blotting. Southern blot analysis demonstrates the presence of multiple Amb a I-related sequences in the ragweed genome. Amb a I is therefore not a single molecule but rather a family of closely related proteins.

Allergens↗
PubMed1991

Sequence polymorphism of Amb a I and Amb a II, the major allergens in Ambrosia artemisiifolia (short ragweed).

Two of the major allergens in the pollen of short ragweed are Amb a I and Amb a II (formerly antigen E and antigen K, respectively). The genes for Amb a I and Amb a II have recently been cloned, and it was shown that Amb a I is a family of proteins with at least three distinct polymorphic family members. This study addresses the number of individual Amb a I and Amb a II family members, the polymorphism in each family member and the expression of these genes in ragweed plants from different geographical locations. This work led to the cloning and characterization of a fourth Amb a I family member, designated Amb a I.4.

Allergens↗
PubMed1991

Serving elders in greatest social and economic need: the challenge to the Elderly Nutrition Program.

The federally funded Elderly Nutrition Program is mandated to serve elders in greatest social and economic need. Yet researchers have concluded that the program underserves certain groups of elders, including those who are socially impaired and homeless and members of ethnic and other minority groups. This paper reviews the issues involved in serving elders in greatest social and economic need and suggests that reaching all of these elders with program services is not only a laudable goal but also, based on innovative models briefly described, is clearly achievable.

Aged↗