Researchers map the complex sugar chains attached to rat serum proteins

A comprehensive study mapped the dynamic glycosylation landscape of rat serum proteins, uncovering sex- and time-dependent variations with clear cell-type specificity. Researchers found distinct cell-type-specific glycosylation patterns: liver-derived proteins carried O-acetylated Neu5Ac, while B cell-derived antibodies (IgG) exclusively utilized Neu5Gc. Female rats exhibited pronounced daily fluctuations in these sugar structures, unlike the stable profiles in males, suggesting regulation by hormonal and circadian cycles. This work provides the first integrated map of glycosylation dynamics in rat serum, establishing a foundation for future studies in comparative glycobiology and biomarker research.

In this study, researchers systematically mapped the complex sugar chains (N-glycans) attached to rat serum proteins, revealing dynamic differences based on sex and time. Researchers identified distinct sialylation signatures that define liver- and immune-derived proteins: liver proteins featured the unique rat-specific modification O-acetylated Neu5Ac, while B cell-derived immune proteins (IgG) exclusively carried N-glycolylneuraminic acid (Neu5Gc). Notably, female rats exhibited pronounced daily fluctuations in these sugar patterns, unlike the stable profiles in males, suggesting strong hormonal regulation.

Glycosylation--the attachment of complex sugar chains (glycans) to proteins--is a fundamental modification crucial for biological processes, and changes in these structures are often associated with development, disease, or aging. Rats serve as essential model organisms, often preferred over mice due to their greater physiological homology to humans and suitability for functional assessments. Previous research highlighted a distinctive feature of rat serum glycans: the unusually high abundance of O-acetylated N-acetylneuraminic acid (Neu5Ac), a modification that is virtually absent in humans. Understanding how physiological factors like sex and daily cycles affect this unique glycosylation pattern is essential for validating rats in studies modeling human pathophysiology.

Researchers employed systematic glycomic (MALDI-TOF-MS) and glycoproteomic (LC-MS/MS) profiling on rat serum collected daily over five consecutive days from male and female rats. To focus on non-IgG proteins, serum albumin and IgG were first depleted from the general serum fraction. A distinguishing feature of the approach was the separate purification and analysis of IgG, which was enzymatically cleaved into its Fab and Fc fragments to obtain cell-type-resolved insights into immune system glycosylation.

The analysis confirmed that the predominant N-glycans are disialo-biantennary complex-type structures, and identified 87 glycoproteins, including protease inhibitors and immune-related proteins like complement C3. A striking finding was the cell-type-specific sialylation profile: liver-derived glycoproteins carried N-acetylneuraminic acid (Neu5Ac), with over half being uniquely O-acetylated (Neu5,9Ac2). In contrast, B cell-derived IgG exclusively carried Neu5Gc and entirely lacked O-acetylation. Crucially, temporal analysis showed that while male rats maintained stable sialylation levels, female rats exhibited significant day-to-day fluctuations, indicating a link to hormonal and circadian rhythms. Furthermore, IgG Fab fragments were approximately three times more sialylated (44% of glycans) than their Fc counterparts (15%).

This comprehensive mapping provides a quantitative framework for future studies using rat models in glycobiology and disease research. The dataset serves as a reference for understanding how hormonal and circadian cycles shape the glycosylation landscape of serum proteins. The observed sex-dependent daily fluctuations in glycosylation patterns highlight a dynamic regulatory link between N-glycosylation and hormonal cycles, which is important for understanding protein stability and function under different physiological states. Elucidating the unique segregation of sialic acid types (Neu5Ac vs. Neu5Gc) between liver and immune tissues underscores the necessity of considering cell-type specificity, informing biomarker discovery and comparative glycoengineering.