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- Creamer, L K, et al.
(author)
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Effect of genetic variation on the tryptic hydrolysis of bovine beta-lactoglobulin A, B, and C
- 2004
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In: Journal of Dairy Science. - 1525-3198. ; 87:12, s. 4023-4032
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Journal article (peer-reviewed)abstract
- The structure, stability, and hydrolysis characteristics of beta-lactoglobulin (LG) A are different from those of either beta-LG B or beta-LG C. Purified samples of these proteins were hydrolyzed with trypsin and the rates of loss of native monomeric beta-LG structure were measured using sodium dodecyl sulfate PAGE. At the same time, the appearance of many individual peptides were identified and followed in time by HPLC, measuring their concentration as a function of solution pH, temperature, protein concentration, and added urea or palmitate. The identity of the peptides was confirmed by liquid chromatography-mass spectrometry. This semiquantitative exploration showed that the rate of hydrolysis was in the order beta-LG A > beta-LG B > beta-LG C under most circumstances, and that 12 of the 18 trypsin-susceptible bonds were cleaved at very similar rates that were governed by the variant type. Consequently, the rate of hydrolysis of the intact protein was related to the overall structural stability of the individual proteins and the accessibility of certain peptide bonds to the enzyme. The hydrolysis of mixtures of 2 or more variants or of denatured beta-LG gave more heterogeneous peptide mixtures.
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- Jimenez, A, et al.
(author)
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Human spermatid-specific thioredoxin-1 (Sptrx-1) is a two-domain protein with oxidizing activity
- 2002
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In: FEBS Letters. - 0014-5793 .- 1873-3468. ; 530:1-3, s. 79-84
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Journal article (peer-reviewed)abstract
- Spermatid-specific thioredoxin-1 (Sptrx-1) is the first member of the thioredoxin family of proteins with a tissue-specific expression pattern, found exclusively in the tail of elongating spermatids and spermatozoa. We describe here further biochemical characterization of human Sptrx-1 protein structure and enzymatic activity. In gel filtration chromatography human Sptrx-1 eluates as a 400 kDa protein consistent with either an oligomeric form, not maintained by intermolecular disulfide bonding, and/or a highly asymmetrical structure. Analysis of circular dichroism spectra of fragments 1-360 and 361-469 and comparison to spectra of full-length Sptrx-1 supports a two-domain organization with a largely unstructured N-terminal domain and a folded thioredoxin-like C-terminal domain. Functionally, Sptrx-1 behaves as an oxidant in vitro when using selenite, but not oxidized glutathione, as electron acceptor. This oxidizing enzymatic activity suggests that Sptrx-1 might govern the stabilization (by disulfide cross-linking) of the different structures in the developing tail of spermatids and spermatozoa.
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