Enzymes and Inhibitors

Lytic Enzymes

<h2>What Are Lytic Enzymes?</h2>
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Lytic enzymes degrade the structural polysaccharide scaffolding of microbial cell walls, enabling cell disruption under mild conditions without mechanical force. Lysozyme (N-acetylmuramide glycanohydrolase) cleaves the beta-1,4 glycosidic bond between N-acetylmuramic acid and N-acetylglucosamine residues in bacterial peptidoglycan, lysing gram-positive bacteria efficiently and gram-negative bacteria when paired with EDTA to destabilize the outer membrane. Lyticase from Arthrobacter luteus hydrolyzes beta-1,3-glucan in yeast cell walls and is the standard reagent for generating spheroplasts from Saccharomyces cerevisiae and other yeast species. Lysostaphin is a glycyl-glycine endopeptidase that specifically cleaves the polyglycine cross-links in Staphylococcus aureus peptidoglycan. Mutanolysin cleaves beta-1,4 linkages in the alternating N-acetylmuramyl-N-acetylglucosamine backbone of gram-positive bacteria, complementing lysozyme for complete lysis of resistant species. Choose the lytic enzyme based on organism type (bacterial vs. yeast/fungal), Gram stain classification, and intended downstream workflow (DNA extraction, protein isolation, or transformation).
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<h2 dir="ltr">What you will find:</h2>
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Zymolyase&trade; Ultra: An elite, high-activity enzyme optimized for rapid fungal cell wall digestion and maximum nucleic acid recovery.
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Flash-Extract Lysis Solution: A streamlined reagent for immediate genomic DNA release from diverse samples, ideal for NGS integration.
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Zymolyase&trade; w/ Storage Buffer: A stabilized formulation designed to maintain long-term enzymatic potency and consistent performance across all assays.
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<h2>How to Choose Lytic Enzymes</h2>
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Gram-positive vs. gram-negative bacteria
Lysozyme alone lyses most gram-positive bacteria efficiently at concentrations of 0.1-1 mg/mL in 10 mM Tris-HCl, pH 8.0, 1 mM EDTA at 37 degrees C for 30-60 minutes. Gram-negative bacteria require EDTA (typically 10-50 mM) to chelate divalent cations and disrupt the outer membrane before lysozyme can access the peptidoglycan. Species with thick, atypical cell walls, such as Micrococcus, Staphylococcus, and Streptococcus, may respond better to lysostaphin, mutanolysin, or mechanical disruption combined with enzymatic lysis.
Yeast and fungal lysis
Lyticase (minimum 2,000 U/mg protein) is the reagent of choice for yeast spheroplast generation and DNA/RNA extraction from Saccharomyces, Candida, Pichia, Kluyveromyces, and related species. A typical protocol uses 10-100 U of lyticase per OD600 unit of yeast cells in 1 M sorbitol buffer at 30 degrees C for 30-60 minutes. Kitalase (from Rhizoctonia solani) combines beta-1,3-glucanase with protease, pectinase, and amylase activity, providing broader lytic activity for resistant fungal species.
DNA purity requirements
For microbiome research and 16S/18S rDNA amplification, lytic enzymes must be certified free of contaminating DNA. Lyticase lots used in microbiome studies are QC-tested by 35-cycle PCR amplification of 16S and 18S rDNA using universal primer sets; confirm this certification with each lot before use in amplicon sequencing workflows.
Downstream compatibility
Most lytic enzymes are inactivated by subsequent SDS treatment or heating to 95 degrees C for 5 minutes, making them compatible with standard nucleic acid extraction kits. Lyticase requires removal before silica column binding; heat inactivation at 70 degrees C for 10 minutes or proteinase K digestion at 55 degrees C for 30 minutes accomplishes this. Confirm inactivation conditions in the manufacturer's protocol before proceeding.
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<h2>Specifications Context</h2>
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Lysozyme activity is measured in units per milligram of protein, where 1 unit is defined as the decrease in absorbance of 0.001 per minute at 450 nm when Micrococcus lysodeikticus cells are used as substrate at pH 6.24 and 25 degrees C. Lyticase activity is expressed as units per milligram protein; 1 unit releases 1 nmol of glucose from yeast cell walls per minute at 30 degrees C. Lysozyme is stable for 12 months at 4 degrees C as a lyophilized powder and should be dissolved in 10 mM sodium acetate, pH 4.5-5.0, for storage in solution. Lytic enzyme combinations with mechanical disruption (bead beating) are standard for complete lysis of difficult samples, including Mycobacterium, spore-forming bacteria, and thick-walled fungi in clinical and microbiome research workflows. For downstream removal of protein from lysates, see <a href="https://mbpinc.net/reagents/molecular-biology/enzymes-and-inhibitors/proteases">proteases,</a> including Proteinase K, for nucleic acid cleanup.
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User Manual

Product Literature

MSDS

Flash-Extract Lysis Solution

AZ-3410 by Azura Genomics: single-tube lysis solution releasing PCR-ready genomic DNA from mouse tails, ear punches, plant leaves, bacteria, and saliva in 8 minutes. Complete workflow under 60 minutes. Available in 100-, 400-, and 2,000-reaction packs. Store at −20°C.

Manufacturer

Flash-Extract Lysis Solution 100 reactions

Flash-Extract Lysis Solution 400 reactions

Proteinase K

<h2>What Is Proteinase K?</h2>
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Proteinase K is a subtilisin-related serine endopeptidase with a molecular weight of 28,930 daltons (28.9 kDa), first isolated from Tritirachium album Limber in 1974, and named for its ability to digest keratin ('K') as its original substrate. The enzyme cleaves peptide bonds at the carboxylic side of aliphatic, aromatic, and basic amino acid residues, making it effective against nearly all protein substrates, including histones, chromatin-bound proteins, cell membrane proteins, and contaminating nucleases (DNases, RNases). Critically, Proteinase K retains full activity in the presence of 1% SDS, 4 M urea, 1% Triton X-100, 0.5% N-lauroylsarcosine, EDTA at 5 mM, and common reducing agents -- conditions under which most other proteases are inactivated. Use Proteinase K in any protocol requiring enzymatic removal of proteins from nucleic acid preparations, including genomic DNA, total RNA, plasmid, phage, and cfDNA extraction from tissues, blood, FFPE samples, bacteria, yeast, and environmental samples.
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<h2 dir="ltr">What you will find:</h2>
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Proteinase K (Lyophilized): A stable, high-purity powder ideal for prolonged storage and tailored buffer preparation.
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Proteinase K Solution: A prepared, stable liquid that reduces preparation time and minimizes pipetting mistakes.
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Proteinase K (G029): A specially refined grade designed for the swift inactivation of RNases and DNases in sensitive procedures.
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<h2>How to Choose Proteinase K</h2>
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Solution vs. lyophilized
Proteinase K is supplied as lyophilized powder (commonly at 30 mAnson units/mg or 20 mg/mL equivalent activity) or as a ready-to-use solution (typically 20 mg/mL in storage buffer). Lyophilized powder offers longer shelf life (up to 2 years at -20 degrees C), cost efficiency for high-volume users, and flexibility in buffer preparation. Solutions are convenient for low-volume use and eliminate weighing errors. Both formats deliver equivalent specific activity when the lot certificate is checked.
Molecular biology grade vs. standard
Molecular biology grade Proteinase K is certified free of detectable endonuclease, exonuclease, DNase, and RNase activities by stringent lot-release assays. Standard grades may carry trace nuclease activity that degrades DNA or RNA during digestion. For any application where nucleic acid integrity matters -- DNA sequencing, NGS library prep, RT-PCR, FISH, or southern blotting -- use only molecular biology grade or equivalent.
Recombinant vs. native
Recombinant Proteinase K (expressed in Pichia pastoris or E. coli) offers freedom from animal-derived material, greater lot-to-lot consistency, and suitability for regulated or GMP-adjacent workflows. Native Proteinase K (isolated from T. album culture) is standard for most research applications. Both must carry DNase/RNase-free certification for nucleic acid use.
Concentration and unit dose
Standard working concentrations are 50-200 microgram/mL for most DNA/RNA extraction protocols in lysis buffer. Higher concentrations of 1-5 mg/mL are used for FFPE tissue deparaffinization and protein digestion, and for prion inactivation studies (where Proteinase K at 10-50 microgram/mL is used to assess proteinase K-resistant PrP accumulation). Proteinase K retains activity for up to 24 months when stored at -20 degrees C in aliquots.
Temperature and pH optimization
Optimal activity is at 20-60 degrees C and pH 7.5-12.0; activity at pH 8.0 and 55 degrees C is used in most nucleic acid extraction protocols. Activity at 37 degrees C is approximately 80% of the 55-60 degrees C optimum. Calcium ions (1 mM Ca2+) stabilize the enzyme structure; EDTA removes calcium and reduces activity, though the enzyme retains sufficient function for nucleic acid prep protocols. Heat-inactivation above 95 degrees C for 5 minutes is complete and does not require chemical inhibitors.
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<h2>Specifications Context</h2>
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Proteinase K specific activity is commonly reported in Anson units (mAnson units/mg): one mAnson unit releases 1 micromol of tyrosine equivalents per minute from hemoglobin at pH 7.5 and 35.5 degrees C. Proteinase K is fully denatured at 95 degrees C in 5 minutes without requiring PMSF or other chemical inhibitors, making heat inactivation the simplest and most reliable option before downstream enzymatic steps such as PCR or reverse transcription. The enzyme is deactivated above 95 degrees C within the first cycle of PCR thermocycling, preventing interference with polymerase activity in protocols that include Proteinase K in the PCR reaction. Proteinase K is a universal reagent in automated nucleic acid extraction platforms, including MagNA Pure, KingFisher, and Hamilton NGS workstations; solution-format Proteinase K is specified in most automated platform kits.&nbsp;
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AZ-3101 by Azura Genomics is a recombinant Proteinase K in lyophilized powder form, molecular biology grade, expressed in Pichia pastoris. RNase-, DNase-, and exonuclease-free with ≤10 pg/mg residual DNA. pH stable 4.0–12.5; active up to 56°C. Available in 100–1000 mg formats.

Proteinase K, 100 mg

Proteinase K, 500 mg

Proteinase K, 1000 mg

Proteinase K Solution

AZ-3201 Proteinase K Solution by Azura Genomics is a ready-to-use 20 mg/ml recombinant serine protease (≥800 U/ml), molecular biology grade, free of nucleases, active at up to 56°C and in urea/SDS. 5 ml, stored at −20°C.