Barley beta-glucosidase: expression during seed germination and maturation and partial amino acid sequences.

Unlike most of the hydrolytic enzymes that participate in endosperm mobilization, beta-glucosidase of barley (Hordeum vulgare) seeds does not increase during germination, even in the presence of exogenously added gibberellic acid. However, the germination process affects the physical properties of beta-glucosidase in terms of charge and apparent molecular weight. Analysis of developing barley grains shows that the enzyme is synthesized two weeks before maturation and is stored in the endosperm of the dry dormant seed. Partial amino acid sequencing of the purified beta-glucosidase demonstrates significant similarity between the barley enzyme and beta-glycosidases that belong to family 1 of glycosyl hydrolases.

The primary structure of DNA binding protein II from the extreme thermophilic bacterium Thermus thermophilus.

The primary structure of DNA binding protein II (DNA bp II) from the extreme thermophilic bacterium Thermus thermophilus has been established by combination of manual and automated techniques. The protein has 95 residues and a molecular mass of 11,843. Comparison of the primary structure with the known sequence data of DNA bp II from Clostridium pasteurineum, Baccillus stearothermophilus, Escherichia coli, Rhizobium meliloti, Anabena, Thermoplasma acidophilum, Pseudomonas aeruginosa and Bacillus caldolyticus reveals a clear homology among these small basic proteins. In particular, two short sequences in the middle and C-terminal part of the proteins (residues N-Gly-Phe-Gly-X-Phe and Pro-X-Thr at positions 46-51 and 63-65, respectively) are completely conserved.

Hormone binding site of the insulin receptor: analysis using photoaffinity-mediated avidin complexing.

A trifunctional reagent was designed which allows derivatization of ligands, particularly peptides and proteins, for subsequent photoaffinity labelling of receptors and specific isolation of the covalent complex or its fragments. B29-(2-nitro-4-azidophenyl)-biocytinyl-insulin (NB-insulin) was synthesized, radioiodinated, and the B26-mono-iodo derivative isolated by HPLC. It was used to photoaffinity label human placental membranes and the purified insulin receptor. Extensive digestion of the covalent insulin-receptor complex with trypsin (EC 3.4.21.4) led to the generation of a fragment of Mr 14,000. Specific complexing with avidin, derivatized avidin or streptavidin could be demonstrated for the photoaffinity labelled alpha-subunit and the 14,000 core fragment. The latter was isolated (approx. 100 pmol from 3-4 placentae) by streptavidin affinity chromatography and HPLC. According to microsequencing based on the known primary structure of the insulin receptor, the N-terminus of the core peptide appears to be Leu20-His21-Glu22-Leu23. We thus conclude: a part of the insulin-binding region of the receptor is located close to the N-terminus of its alpha-subunit in a remarkably stable domain of the sequence 20--(approx.) 120.