Crystal size, a key character of lactose crystallization affecting microstructure, surface chemistry and reconstitution of milk powder.

Lactose crystallization during storage deteriorates reconstitution performance of milk powders, but the relationship between lactose crystallization and reconstitution is inexplicit. The objective of this study is to characterize crystalline lactose in the context of formulation and elucidate the complex relationship between lactose crystallization and powder functionality. Lactose in Skim Milk Powder (SMP), Whole Milk Powder (WMP) and Fat-Filled Milk Powder (FFMP) stored under 23 %, 53 % and 75 % Relative Humidity (RH) at 25  â„ƒ for four months was compared. Lactose, surface chemistry and microstructure of FFMP stored at 25 â„ƒ and 40 â„ƒ at 23 % to 75 % RH for four months were also analyzed and interpreted. At the same RH, FFMP crystallized in the same pattern as WMP. At 53 % RH, FFMP and WMP differentiated from SMP in terms of lactose morphology as well as the ratio between anhydrous α-lactose and anhydrous ß-lactose. Lactose remained amorphous at 23 % RH, crystallized predominantly to α/ß-lactose (1:4) at 40 to 58 % RH and to α-lactose monohydrate at 75 % RH. The crystallinity index was similar for all powders containing crystalline lactose. The estimated crystallite size increased from approx. 0.1 to 20 µm with increasing RH and temperature. When amorphous lactose crystallized into crystals below approx. 0.1 µm at 25 °C and 43 % RH, the microstructure and surface lipid were comparable to that of the reference powder. This powder reconstituted into a stable suspension system comparable to that of reference (well performing) powders. These results demonstrate that crystallite size is the key property linking lactose crystallization and reconstitution. Our finding thus indicates limiting crystallite size is important for maintaining desired product quality.

Corneal Dystrophy Mutations Drive Pathogenesis by Targeting TGFBIp Stability and Solubility in a Latent Amyloid-forming Domain.

Numerous mutations in the corneal protein TGFBIp lead to opaque extracellular deposits and corneal dystrophies (CDs). Here we elucidate the molecular origins underlying TGFBIp's mutation-induced increase in aggregation propensity through comprehensive biophysical and bioinformatic analyses of mutations associated with every major subtype of TGFBIp-linked CDs including lattice corneal dystrophy (LCD) and three subtypes of granular corneal dystrophy (GCD 1-3). LCD mutations at buried positions in the C-terminal Fas1-4 domain lead to decreased stability. GCD variants show biophysical profiles distinct from those of LCD mutations. GCD 1 and 3 mutations reduce solubility rather than stability. Half of the 50 positions within Fas1-4 most sensitive to mutation are associated with at least one known disease-causing mutation, including 10 of the top 11 positions. Thus, TGFBIp aggregation is driven by mutations that despite their physico-chemical diversity target either the stability or solubility of Fas1-4 in predictable ways, suggesting straightforward general therapeutic strategies.

How Glycosaminoglycans Promote Fibrillation of Salmon Calcitonin.

Glycosaminoglycans (GAGs) bind all known amyloid plaques and help store protein hormones in (acidic) granular vesicles, but the molecular mechanisms underlying these important effects are unclear. Here we investigate GAG interactions with the peptide hormone salmon calcitonin (sCT). GAGs induce fast sCT fibrillation at acidic pH and only bind monomeric sCT at acidic pH, inducing sCT helicity. Increasing GAG sulfation expands the pH range for binding. Heparin, the most highly sulfated GAG, binds sCT in the pH interval 3-7. Small angle x-ray scattering indicates that sCT monomers densely decorate and pack single heparin chains, possibly via hydrophobic patches on helical sCT. sCT fibrillates without GAGs, but heparin binding accelerates the process by decreasing the otherwise long fibrillation lag times at low pH and accelerates fibril growth rates at neutral pH. sCT·heparin complexes form ß-sheet-rich heparin-covered fibrils. Solid-state NMR reveals that heparin does not alter the sCT fibrillary core around Lys(11) but makes changes to Val(8) on the exterior side of the ß-strand, possibly through contacts to Lys(18) Thus GAGs significantly modulate sCT fibrillation in a pH-dependent manner by interacting with both monomeric and aggregated sCT.

A Complex Dance: The Importance of Glycosaminoglycans and Zinc in the Aggregation of Human Prolactin.

The zinc binding hormone pituitary human prolactin (hPRL) is stored in secretory granules of specialized cells in an aggregated form. Glycosaminoglycans (GAGs) are anionic polysaccharides commonly associated with secretory granules, indicating their involvement in granule formation. Here we, for the first time, study the impact of GAGs in combination with Zn(2+) on the reversible hPRL aggregation across the pH range of 7.4-5.5. Zn(2+) alone causes hPRL aggregation at pH 7.4, while aggregation between pH 7.4 and 5.5 requires both Zn(2+) and GAGs. GAGs alone cause hPRL aggregation below pH 5.5. Comprehensive thermal stability investigations show that hPRL is particularly destabilized toward thermal denaturation at pH 5.5 and that GAGs increasingly destabilize hPRL at decreasing pH values. We propose that Zn(2+) causes hPRL aggregation through low-affinity Zn(2+) binding sites on hPRL with GAGs facilitating Zn(2+) binding by neutralizing repulsive positive charges of hPRL in the acidic environments of the TGN and mature secretory granules. In a manner independent of the aggregation-causing agent(s), the different hPRL aggregates show very similar secondary structure and amorphous morphology. We speculate that this may be a recognizable sorting signal in the formation of hPRL granular vesicles.