N-glycosylation is one of the most important posttranslational modifications of proteins. It plays important roles in the biogenesis and functions of proteins by influencing their folding, intracellular localization, stability and solubility. N-glycans are synthesized by glycosyltransferases, a complex group of ubiquitous enzymes that occur in most kingdoms of life. A growing body of evidence shows that N-glycans may influence processing and functions of glycosyltransferases, including their secretion, stability and substrate/acceptor affinity. Changes in these properties may have a profound impact on glycosyltransferase activity. Indeed, some glycosyltransferases have to be glycosylated themselves for full activity. N-glycans and glycosyltransferases play roles in the pathogenesis of many diseases (including cancers), so studies on glycosyltransferases may contribute to the development of new therapy methods and novel glycoengineered enzymes with improved properties. In this review, we focus on the role of N-glycosylation in the activity of glycosyltransferases and attempt to summarize all available data about this phenomenon.

Rare polyagglutinable NOR erythrocytes contain three unique globoside (Gb4Cer) derivatives, NOR1, NOR_int, and NOR2, in which Gal(α1–4), GalNAc(β1–3)Gal(α1–4), and Gal(α1–4)GalNAc(β1–3)Gal(α1–4), respectively, are linked to the terminal GalNAc residue of Gb4Cer. NOR1 and NOR2, which both terminate with a Gal(α1–4)GalNAc- sequence, react with anti-NOR antibodies commonly present in human sera. While searching for an enzyme responsible for the biosynthesis of Gal(α1–4)GalNAc, we identified a mutation in the A4GALT gene encoding Gb3/CD77 synthase (α1,4-galactosyltransferase). Fourteen NOR-positive donors were heterozygous for the C>G mutation at position 631 of the open reading frame of the A4GALT gene, whereas 495 NOR-negative donors were homozygous for C at this position. The enzyme encoded by the mutated gene contains glutamic acid instead of glutamine at position 211 (substitution Q211E). To determine whether this mutation could change the enzyme specificity, we transfected a teratocarcinoma cell line (2102Ep) with vectors encoding the consensus Gb3/CD77 synthase and Gb3/CD77 synthase with Glu at position 211. The cellular glycolipids produced by these cells were analyzed by flow cytometry, high-performance thin-layer chromatography, enzymatic degradation, and MALDI-TOF mass spectrometry. Cells transfected with either vector expressed the P1 blood group antigen, which was absent from untransfected cells. Cells transfected with the vector encoding the Gb3/CD77 synthase with Glu at position 211 expressed both P1 and NOR antigens. Collectively, these results suggest that the C631G mutation alters the acceptor specificity of Gb3/CD77 synthase, rendering it able to catalyze synthesis of the Gal(α1–4)Gal and Gal(α1–4)GalNAc moieties.

Serum of animals belonging to the Camelidae family (camels and llamas) contains fully active antibodies that are naturally devoid of light chains. Variable domains derived from heavy chain antibodies (hcAb) called VHHs or nanobodies™ can bind antigens as effectively as full-length antibodies and are easy to clone and express. Because of their potential, VHHs are being intensively studied as potential therapeutic, diagnostic and imaging tools. The paper reviews the molecular background of heavy chain antibodies and describes methods of obtaining recombinant fragments of heavy chain antibodies as well as their therapeutic, diagnostic and other applications.