In enzyme immobilization by covalent bonding, the enzyme molecules are coupled to the support via which type of amino acid residues and what kind of support material is typically used?

Introduction:
Covalent immobilization forms stable, permanent bonds between an enzyme and a solid support. The goal is to retain catalytic activity while preventing enzyme leaching during operation. Choosing which residues participate and the nature of the support are central to performance, activity retention, and reusability.

Given Data / Assumptions:

• Covalent attachment creates chemical bonds between enzyme side chains and activated groups on a support.
• Active site residues are essential for catalysis and should be preserved.
• Industrial supports are typically water-insoluble and functionalized (for example, epoxy, aldehyde, carbodiimide-activated carboxyl, cyanogen bromide activated).

Concept / Approach:
To avoid activity loss, immobilization chemistry targets nonessential surface residues such as lysine epsilon-amines, cysteine thiols away from the catalytic pocket, or carboxyl groups not involved in catalysis. Water-insoluble supports (for example, cross-linked agarose, silica, polymethacrylate) provide mechanical strength, easy separation, and low pressure drop in packed beds. Functionalization allows selective, mild coupling conditions that minimize enzyme denaturation.

Step-by-Step Solution:

Verification / Alternative check:
Activity assays before and after immobilization, along with kinetic parameters (apparent Km and Vmax), demonstrate retained function. Reduced protein in effluent across cycles confirms negligible leaching characteristic of covalent bonding.

Why Other Options Are Wrong:

• Essential residues on water-insoluble supports: risks direct active site modification and inactivation.
• Water-soluble supports: complicate separation and promote leaching in flow reactors.
• Any scheme focusing on essential residues is unsafe for activity retention.

Common Pitfalls:
Over-activation causing multipoint attachment near the active site can restrict conformational mobility. High temperatures or extreme pH during coupling may denature the enzyme. Proper spacer arms often improve accessibility and activity.

Final Answer:
nonessential amino acids residues to water insoluble, functional supports