Mutations at the dimer, hexamer, and receptor-binding surfaces of insulin independently affect insulin-insulin and insulin-receptor interactions
- Harvard Medical School, Boston, MA (United States)
Mutagenesis of the dimer- and hexamer-forming surfaces of insulin yields analogues with reduced tendencies to aggregate and dramatically altered pharmacokinetic properties. The authors recently showed that one such analogue, HisB1- {yields} Asp, ProB28 {yields} Lys, LysB29 {yields} Pro human insulin (DKP-insulin), has enhanced affinity for the insulin receptor and is useful for studying the structure of the insulin monomer under physiologic solvent conditions. DKP-insulin retains native secondary and tertiary structure in solution and may therefore provide an appropriate baseline for further studies of related analogues containing additional substitutions within the receptor-binding surface of insulin. To test this, they prepared a family of DKP analogues having potency-altering substitutions at the B24 and B25 positions using a streamlined approach to enzymatic semisynthesis which negates the need for amino-group protection. For comparison, similar analogues of native human insulin were prepared by standard semisynthetic methods. The DKP analogues show a reduced tendency to self-associate, as indicated by {sup 1}H-NMR resonance line widths. Such 'template independence' reflects an absence of functional interactions between the B24 and B25 sites and additional substitutions in DKP-insulin and demonstrates that mutations in discrete surfaces of insulin have independent effects on protein structure and function. In particular, the respective receptor-recognition (PheB24, PheB25), hexamer-forming (HisB10), and dimer-forming (ProB28, LysB29) surfaces of insulin may be regarding as independent targets for protein design. DKP-insulin provides an appropriate biophysical model for defining structure-function relationships in a monomeric template.
- OSTI ID:
- 5559639
- Journal Information:
- Biochemistry; (United States), Vol. 31:6; ISSN 0006-2960
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
INSULIN
NUCLEAR MAGNETIC RESONANCE
RECEPTORS
CONFORMATIONAL CHANGES
ASPARTIC ACID
DICHROISM
HISTIDINE
LIQUID COLUMN CHROMATOGRAPHY
LYSINE
MUTAGENESIS
MUTATIONS
AMINO ACIDS
AZOLES
CARBOXYLIC ACIDS
CHROMATOGRAPHY
HETEROCYCLIC ACIDS
HETEROCYCLIC COMPOUNDS
HORMONES
IMIDAZOLES
MAGNETIC RESONANCE
MEMBRANE PROTEINS
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
PEPTIDE HORMONES
PROTEINS
RESONANCE
SEPARATION PROCESSES
550201* - Biochemistry- Tracer Techniques