Study Guide 19: Carbohydrate Technologies

Glycan

Complex oligo/polysaccharide portion of a glycoconjugate

N-linked glycans

Attached to the amide nitrogen of an asparagine (Asn) side chain

O-linked glycans

Attached to the hydroxyl oxygen of serine (Ser) or threonine (Thr) side chains

Biological roles of glycans

- cell-cell recognition and signaling

- pathogen interactions and infection

- protein folding and stability

Glycoconjugates

glycoproteins and glycolipids

How to identify glycosylation sites

UniProt (PTM/Processing section)

Glycosyltransferases

Catalyze the transfer of a sugar moiety from an activated donor (nucleotide-activated sugar) to an acceptor molecule, forming a glycosidic bond

Glycan composition determines blood types

It depends on the type of glycosyltransferases (genetics)

Type A blood

A antigens and anti-B antibodies

Type B blood

B antigens and anti-A antibodies

Type AB blood

A and B antigens, no antibodies

Type O blood

no antigens, A and B antibodies

Sialic acid

Modified carbohydrate linked to proteins to form N- or O- linked glycans on glycoproteins

Sialic Acid Glycans

upregulated on the surface of cancer cells, playing a crucial role in immune modulation and tumor immune evasion. The surface of cancer cells has elevated sialic acid glycans

Sialic acid and cancer

Sialic acid glycans interact with Siglec receptors, inhibiting immune cells. Removal of sialic acids by an enzyme antibody (sialidase- glycosidase) conjugate stops the interaction of sialylated glycans and NK-inhibitory Siglec receptors.

Protonation state

At physiological pH, it's protonated. (Due to the pka values, most carbohydrates hydroxyl groups will be protonated at physiological pH.) *Weak acids

Reducing vs non-reducing carbohydrates

Reducing has a hemiacetal (aldose- OR2) or hemiketal (ketose- OR3) group. Non-reducing has acetal (OR, H) or ketal linkage (OR, OR)

Benedict's test

Colorimetric assays to test for reducing vs non-reducing sugars. Blue indicates sugar is not reducing. Red indicates large amount of reducing sugar. Benedict's reagent is an alkaline solution containing copper(II) sulfate (oxidizing agent/source of ), sodium carbonate (basic medium), and sodium citrate (complexing agent).

Carbohydrate analysis

Difficult because they all have somewhat similar physical and chemical characteristics. They also lack strong chromophores and don't absorb significantly in the UV-visible range (200-400 nm).

Eastern Blotting

post-translational modifications on proteins. Glycoproteins are separated by SDS-PAGE and transferred to a membrane. The membrane is probed with lectins (which selectively recognizes carbohydrates) instead of antibodies.

Circular Dichroism

Studies protein secondary and tertiary structures.

Endoglycosidases

Catalyze the hydrolysis of glycosidic linkages within oligosaccharide chains.

Exoglycosidases

Catalyzes the hydrolysis of single sugar residues sequentially from the non-reducing end of oligosaccharide chains (Caroline)

HPLC

High-performance liquid chromatography is used to separate, identify, and quantify mixtures

Mass Spectrometry & NMR

Based on the ionization of compounds when they are bombarded with a beam of electrons to form ions. NMR offers "fingerprinting" all types of carbohydrates

What are some carbohydrate features of sialic acid? Is it D or L? Is it an Aldose or Ketose? Where are the modifications? What ring?