The amino-acid sequence and three-dimensional structure of equine serum albumin have been determined. The amino-acid sequence was deduced from cDNA isolated from equine liver. Comparisons of the primary structure of equine serum albumin with human serum albumin and bovine serum albumin reveal 76.1 % and 73.9% sequence identity, respectively. The three-dimensional structure was determined crystallographically by the molecular-replacement method using molecular coordinates from the previously determined structure of human serum albumin, to a resolution of 0.27 nm. In accordance with the primary structure, the three-dimensional structures are highly conserved. There is a root-mean-square difference between a-carbons of the two structures of 0.201 nm. The association constants (Ka) for the binding of 2,3,5-triiodobenzoic acid were determined by ultrafiltration methods for equine and human serum albumins to be approximately 1WM-l and 105M-', respectively. Crystallographic studies of equine serum albumin reveal two binding sites for 2,3,5-triiodobenzoic acid identical with those previously reported for human serum albumin which are located within subdomains IIA and IIIA. Details and comparisons of the binding chemistry are discussed.As the most abundant protein of the circulatory system albumin functions as a multipurpose transport molecule and is a principal contributor to colloid osmotic blood pressure.

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ABSIRA4CA derivative of bacteriophage X, X 21, has been constructed and used for the cloning of Escherichia coli DNA fragments carrying promoters. Phaage X 21 lacks the lac promoter-operator and can accept DNA fragments up to 9.8 kilobases in size at a unique HindIII restriction endonuclease site adjacent to lacZ -Recombinant phage that carry promoters are readily identified by their expression of lacZ Lysogens of these phages in strains harboring a deletion of the chromosomal lac operon are capable of growth on lactose as sole carbon source and can be used to study some of the regulatory signals that act upon the cloned promoter. In Promoter regions and adjacent sequences are of prime importance in understanding the molecular mechanisms of gene regulation. The fundamental problem of recognizing and defining promoter sites has been approached'through the isolation of up-or down-promoter mutants and the analysis of deletions or operon fusions (1-5). However, for those operons that specify products not readily' detectable, it is difficult to measure promoter activity or to select promoter mutants. This applies to a large group of essential genes, such as those for ribosomal proteins (r-proteins) and rRNA' whose study would be more expeditious were it not for the tedious procedures necessary for quantitation of their gene products. One way of circumventing these difficulties is to devise a means of fusing promoters to promoterless structural genes whose products are conveniently measured, such as lacZ (3). With the available increasingly detailed catalogue of promoter regions determined with restriction enzymes, it is now possible to extend this approach by in vitro construction of cloning vehicles. These would be of general use in the direct isolation of promoter fragments (6-8). MATERIALS AND METHODSBacterial and Phage Strains. The Escherichia coli K-12 strains were from our collection: NF1531 F-minA -minBthi-/pEH1 (9), JF1598 HfrH A(lac)X74 thi-relAl, JF1708 F-thr-A(lac)X74 his pyrE-argHjfucr rpsL31 this-, JF2171 F-A(lac)X74 trpE9851 rpsL.31 rho-4 thi-(10). Bacteriophage A p123(209) was obtained from M. Casadaban (3), A 540 from N. Murray (11), and X drifdl8 from J. Kirschbaum (12).Media, Phage Growth and Purification, and DNA Preparation. M9 minimal medium (13) was used and supplemented with 50Ug of L amino acids per ml and 0.4% carbon source. NZ amine (Humko, Norwich, NY) medium described by Blattner et al. (14) was used for growth of phage. Cells for phage A infection were grown in L broth (15) containing 0.4% maltose.5-Bromo-4-chloro-3-indolyl-f3-D-galactosidase (XG) (Sigma) was used at a concentration of 40 ,g/ml.A phage carrying the S7 allele were grown and purified as described (16). All other X phage were grown by the "PDS" method and purified as described by Blattner et al. (14). Phage DNA was purified as described (17) 20 Al of NaDodSO4 sample buffer (9). Extracts were prepared by boiling for 5 min and were then resolved on one-dimensional 10-20% polyacrylamide/NaDodSO4 gradient slab gels according t...

The major histocompatibility complex of the rat (RT1 complex) encodes two sets of class II molecules referred to as RT1.B and RT1.D. The complete structure of the RT1.D alpha u chain of the diabetes prone BB rat was determined by the isolation and characterization of a full size cDNA. Comparisons of the nucleotide and protein sequences of RT1.D alpha with the analogous molecules, H-2 I-E alpha and HLA DR alpha, revealed that these alpha chains have been highly conserved during evolution. Southern blot analysis indicated an association of the RT1 haplotypes, 'u' and 'l', with Bam H1 DNA bands of 9.8 kb and 11.7 kb, respectively. The BB rat develops insulin dependent diabetes as an autoimmune abnormality. Accumulating evidence suggests a cellular mediated etiology and the involvement of class II molecules. The steady state levels of RT1.D alpha mRNA were measured in splenic lymphocytes of diabetes prone BB rats and age matched histocompatible normal nondiabetic WF rats by a RNase protection assay. Compared to WF rats, elevated transcripts of RT1.D alpha were found in lymphocytes of young BB rats (approximately 4x and approximately 2.5x greater at 20-40 and 40-75 d, respectively). In lymphocytes of older diabetic and nondiabetic BB rats (greater than 75 d) the levels of RT1.D alpha mRNA were lower than in the young BB rats and were found at the WF control levels. The increased steady state RT1.D alpha mRNA levels in the young BB rats may reflect differences in the proportion of splenic lymphocytes expressing this gene (activated lymphocytes), and thus differences in splenic lymphocyte populations. The steady state RT1.D alpha mRNA levels in lymphocytes of the normal rats were found to be relatively similar at all ages examined. The increased class II gene transcripts found in lymphocytes of young BB rats indicates that they possess a highly activated immune system.